Bone Marrow Response Research Articles

Mapping Bone Marrow Response in the Vertebral Column by Positron Emission Tomography Following Radiotherapy and Erlotinib Therapy of Lung Cancer

To map functional bone marrow (BM) by 2-deoxy-2-[18F]fluoro-D-glucose ([18F]FDG) positron emission tomography (PET) in the vertebral column of lung cancer patients prior to, during, and after treatment. Moreover, to identify radiation- and erlotinib-induced changes in the BM. Twenty-six patients with advanced non-small cell lung cancer, receiving radiotherapy (RT) alone or concomitantly with erlotinib, were examined by [18F]FDG PET before, during, and after treatment. A total of 61 [18F]FDG PET scans were analyzed. Vertebral column BM [18F]FDG standardized uptake value normalized to the liver (SUVBMLR) was used as uptake measure. Wilcoxon signed-rank test was used to assess changes in BM uptake of [18F]FDG between sessions. Effects of erlotinib on the BM activity during and aftertreatment were assessed using Mann-Whitney U test. A homogeneous uptake of [18F]FDG was observed within the vertebral column prior to treatment. Mean SUVBMLR (± S.E.M) in the body of thoracic vertebrae receiving a total RT dose of 10Gy or higher was 0.64 ± 0.01, 0.56 ± 0.01, and 0.59 ± 0.01 at pre-, mid-, and post-therapy, respectively. A significant reduction in the mean SUVBMLR was observed from pre- to both mid- and post-therapy (p < 0.05). Mean SUVBMLR was significantly higher at post-therapy compared to mid-therapy for patients receiving erlotinib in addition to RT (p < 0.05). RT reduces BM [18F]FDG uptake in the vertebral column, especially in the high-dose region. Concomitant erlotinib may stimulate a recovery in BM [18F]FDG uptake from mid- to post-therapy. NCT02714530. Registered 10 September 2015.

Risk prediction based on post induction bone marrow response and genomic profile: A new way to stratify stage M neuroblastoma patients?

10550Background: So far, and apart from minor adaptations, high-risk neuroblastoma patients have been treated uniformly by not taking biological features of the tumor into consideration. To mend th...

Aging alters the immunological response to ischemic stroke

The peripheral immune system plays a critical role in aging and in the response to brain injury. Emerging data suggest inflammatory responses are exacerbated in older animals following ischemic stroke; however, our understanding of these age-related changes is poor. In this work, we demonstrate marked differences in the composition of circulating and infiltrating leukocytes recruited to the ischemic brain of old male mice after stroke compared to young male mice. Blood neutrophilia and neutrophil invasion into the brain were increased in aged animals. Relative to infiltrating monocyte populations, brain-invading neutrophils had reduced phagocytic potential, and produced higher levels of reactive oxygen species and extracellular matrix-degrading enzymes (i.e., MMP-9), which were further exacerbated with age. Hemorrhagic transformation was more pronounced in aged versus young mice relative to infarct size. High numbers of myeloperoxidase-positive neutrophils were found in postmortem human brain samples of old (> 71 years) acute ischemic stroke subjects compared to non-ischemic controls. Many of these neutrophils were found in the brain parenchyma. A large proportion of these neutrophils expressed MMP-9 and positively correlated with hemorrhage and hyperemia. MMP-9 expression and hemorrhagic transformation after stroke increased with age. These changes in the myeloid response to stroke with age led us to hypothesize that the bone marrow response to stroke is altered with age, which could be important for the development of effective therapies targeting the immune response. We generated heterochronic bone marrow chimeras as a tool to determine the contribution of peripheral immune senescence to age- and stroke-induced inflammation. Old hosts that received young bone marrow (i.e., Young → Old) had attenuation of age-related reductions in bFGF and VEGF and showed improved locomotor activity and gait dynamics compared to isochronic (Old → Old) controls. Microglia in young heterochronic mice (Old → Young) developed a senescent-like phenotype. After stroke, aged animals reconstituted with young marrow had reduced behavioral deficits compared to isochronic controls, and had significantly fewer brain-infiltrating neutrophils. Increased rates of hemorrhagic transformation were seen in young mice reconstituted with aged bone marrow. This work suggests that age alters the immunological response to stroke, and that this can be reversed by manipulation of the peripheral immune cells in the bone marrow.

Sphingosine-1-phospate receptor 4 regulates passive systemic anaphylaxis in mice but is dispensable for canonical IgE-mediated responses in mast cells

Abstract Mast cells are key players in the development of inflammatory allergic reactions. Cross-linking of the high-affinity receptor for IgE (FcɛRI) on mast cells leads to the generation and secretion of the sphingolipid mediator, sphingosine-1-phosphate (S1P) which can in turn activate its G-protein coupled receptors on mast cells. Previous reports have identified the expression of two of the five receptors for S1P on mast cells, S1P1 and S1P2, with functions in IgE-receptor-mediated chemotaxis and degranulation, respectively. Here, we show that cultured human and mouse mast cells also express abundant message for the S1P4 receptor and demonstrate its presence at the plasma membrane in the human mast cell line LAD2. Genetic deletion of S1pr4 resulted in exacerbation of passive systemic anaphylaxis to IgE/anti-IgE in mice. However, exhaustive characterization of IgE-mediated responses in S1P4-deficient bone marrow and peritoneal mouse mast cells indicated that this receptor is dispensable for normal in vitro mast cell growth and responses to IgE-mediated activation including degranulation, cytokine/chemokine release and chemotaxis. Thus, we provide evidence that S1P4 modulates mast cell-mediated anaphylaxis in vivo in an unexpected manner that does not involve regulation of mast cell responsiveness to IgE stimulation.

IMMATURE PLATELET FRACTION DI DEMAM DENGUE DAN DEMAM BERDARAH DENGUE (Immature Platelet Fraction in Dengue Fever and Dengue Hemorrhagic Fever)

Thrombocytopenia is a hematological abnormality found in the majority of Dengue Virus Infection cases with manifestations suchas Dengue Fever (DF) and Dengue Hemorrhagic Fever (DHF). Bone marrow response to the decrease in platelets is by increasingthrombopoiesis which can be identified by Immature Platelet Fraction (IPF) examination as an indirect indicator of bone marrow responseto thrombocytopenia. The examination of IPF in venous blood was performed on 29 subjects who met the 1997 WHO criteria, carriedout from January until August 2012. The EDTA blood samples were examined twice, on the day of their admittance and two days later,based on a flowcytometry principle using Sysmex XE-2100. The IPF was derived from the immature platelet ratio against the total numberof platelets (IPF %). The test results were statistically analyzed by using SPSS 20. It was found, that IPF in DHF compared between thefirst and the third day of their admittance was statistically significantly different with p = 0.033 compared to DF with p = 0.444. ThePearson's correlation showed an inverse correlation between IPF and platelets with r = -0.675 and p = 0.01. The statistical analysisrevealed a significant difference in IPF between moderate- and mild-thrombocytopenia on the first and third day of their admittance withp = 0.014 and 0.001, respectively. Based on this study it can be concluded that IPF can be used to indicate the bone marrow response inboth DF and DHF related to thrombocytopenia.

Innate Immunity in the Persistent Inflammation, Immunosuppression, and Catabolism Syndrome and Its Implications for Therapy

Clinical and technological advances promoting early hemorrhage control and physiologic resuscitation as well as early diagnosis and optimal treatment of sepsis have significantly decreased in-hospital mortality for many critically ill patient populations. However, a substantial proportion of severe trauma and sepsis survivors will develop protracted organ dysfunction termed chronic critical illness (CCI), defined as ≥14 days requiring intensive care unit (ICU) resources with ongoing organ dysfunction. A subset of CCI patients will develop the persistent inflammation, immunosuppression, and catabolism syndrome (PICS), and these individuals are predisposed to a poor quality of life and indolent death. We propose that CCI and PICS after trauma or sepsis are the result of an inappropriate bone marrow response characterized by the generation of dysfunctional myeloid populations at the expense of lympho- and erythropoiesis. This review describes similarities among CCI/PICS phenotypes in sepsis, cancer, and aging and reviews the role of aberrant myelopoiesis in the pathophysiology of CCI and PICS. In addition, we characterize pathogen recognition, the interface between innate and adaptive immune systems, and therapeutic approaches including immune modulators, gut microbiota support, and nutritional and exercise therapy. Finally, we discuss the future of diagnostic and prognostic approaches guided by machine and deep-learning models trained and validated on big data to identify patients for whom these approaches will yield the greatest benefits. A deeper understanding of the pathophysiology of CCI and PICS and continued investigation into novel therapies harbor the potential to improve the current dismal long-term outcomes for critically ill post-injury and post-infection patients.

Prognostic significance of leukopenia during the induction phase in adult B cell acute lymphoblastic leukemia.

The association between chemotherapy-induced leukopenia and clinical outcome has been reported for several types of cancer. The objective of the current study was to evaluate the association of chemotherapy-induced leukopenia during the induction phase with the clinical outcome of adult B cell acute lymphoblastic leukemia (B-ALL). Fifty-one cases of B-ALL, age ≥14 years, were reviewed. The variables under consideration included age, sex, the initial white blood cell (WBC) count (WBC-0), as well as the WBC counts on days 8 (WBC-8), 15 (WBC-15), and 22 (WBC-22) during induction therapy, early bone marrow responses on day 15 during induction therapy, immunophenotype, and cytogenetics. Univariate analysis revealed that WBC-15 ≥0.40×109/L was significantly associated with inferior event-free survival (EFS) (hazard ratio [HR]=2.95, P=0.004) and overall survival (OS) (HR=2.92, P=0.015). On multivariate analysis, high WBC-15 (≥0.40×109/L) remained an independent prognostic factor for EFS (HR=3.29, P=0.014) and OS (HR=3.29, P=0.038). Our results suggested that WBC-15 may contribute to refinements in the current risk stratification algorithms for adult B-ALL.

New bioactive glass scaffolds with exceptional qualities for bone tissue regeneration: response of osteoblasts and osteoclasts

Tissue regeneration is a significantly improved alternative to tissue replacement by implants. It requires porous bioscaffolds for the restoration of natural tissue rather than relying on bio-inactive, often metallic implants. Recently, we developed technology for fabricating novel, nano-macroporous bioactive ‘tailored amorphous multi-porous (TAMP)’ hard tissue scaffolds using a 70 mol% SiO2–30 mol% CaO model composition. The TAMP silicate scaffolds, fabricated by a modified sol-gel process, have shown excellent biocompatibility via the rapid formation of hydroxyapatite in biological fluids as well as in early tests with bone forming cells. Here we report an in depth investigation of the response of MC3T3-E1 pre-osteoblast cells and bone marrow derived (BMD) osteoclasts to these TAMP scaffolds. Light and electron microscopic imaging, gene and protein expression, and enzyme activity analyses demonstrate that MC3T3-E1 pre-osteoblasts adhere, proliferate, colonize, and differentiate on and inside the bioactive TAMP scaffolds. Additionally, BMD precursor cells mature into active osteoclasts and remodel the scaffold, highlighting the exceptional qualities of this novel scaffold material for bone tissue regeneration.

Patients over 40 years old with precursor T-cell lymphoblastic lymphoma have different prognostic factors comparing to the youngers

This study aimed to analyze the clinical characteristics and prognostic factors of patients, divided into over 40-year-old group or not, with precursor T-cell lymphoblastic lymphoma (Pre-T-LBL). Based on the retrospective analysis of the clinical data of 59 patients with Pre-T-LBL during the period from December 2010 to December 2015, albumin level, anemia, pleural or pericardial effusion, protocol, therapy response, mediastinal mass, lactate dehydrogenase (LDH), and international prognostic index (IPI) or age-adjusted international prognostic index (aaIPI) were summarized. For patients aged <40 years, factors correlating with poor progression-free survival (PFS) were pleural or pericardial effusion, regimen, albumin level and therapy response. Pleural or pericardial effusion, aaIPI score, regimen, LDH increased, albumin level, therapy response and mediastinal mass were all related with poor overall survival (OS). In the patients aged ≥40 years, only anemia associated with PFS. However, anemia, involvement of bone marrow and therapeutic response were all related with poor OS. In conclusion, the patients with Pre-T-LBL are characterized by a low incidence and bad prognosis. Different prognostic factors can be discovered for patients over 40-year-old with Pre-T-LBL comparing to the youngers. New prognostic evaluation factors should be explored for patients ≥40 years old.

Initial response of human bone marrow–derived stem cells after contact with ultrahigh-molecular-weight polyethylene (UHMWPE) material: An in vitro study on cell viability and interleukin-6 expression

Introduction:Ultrahigh-molecular-weight polyethylene (UHMWPE) is a thermoplastic polymer useful in biomaterial applications, especially in orthopedic field. Yet, little is known concerning its initial effect on human bone marrow stem cells (hBMSCs) after implantation.Materials and Methods:A cytotoxicity analysis was performed with a 3-(4,5-dimethylthiazol 2-yl)-2,5-diphenyltetrazolium assay after 24, 48, and 72h of incubation of hBMSC culture. Expression of interleukin-6 (IL-6) was measured using enzyme-linked immunosorbent assay. Cell viability was measured with Inhibitory concentration 50% (IC50) formula.Results:All treatment groups showed a cell viability of >50% ranging from 78% to >100%. Lower expression of IL-6 of hBMSC compared to control group was found in 48h of incubation period.Conclusion:hBMSC showed high cell viability after initial contact with UHMWPE material. Modulation of IL-6 expression was present at the initial stage as a response to foreign material.

Modic type 1 change is an autoimmune response that requires a proinflammatory milieu provided by the ‘Modic disc’

Background ContextModic changes (MCs) are magnetic resonance imaging (MRI) evidence of inflammatory and fibrotic vertebral bone marrow lesions that associate with adjacent disc degeneration and end plate damage. Although MC etiology is uncertain, historical data suggest a linkage to an autoimmune response of bone marrow triggered by the nucleus pulposus (NP). PurposeThe aim of this study was to test whether bone marrow has an autoimmune response to NP cells that is amplified by an inflammatory milieu and ultimately leads to MC development in vivo. We hypothesized that an inflammatory co-stimulus is required for bone marrow/NP crosstalk to stimulate MC. Study DesignThis is an in-vitro cell co-culture study plus in-vivo experiments in rat caudal vertebrae. MethodsIn in-vitro study, bone marrow mononuclear cells (BMNCs) and NP cells (NPCs) from rats were co-cultured with and without interleukin (IL)-1α stimulation. Cell viability (n=3) of BMNCs and NPCs and gene expression (n=7) were analyzed. In in-vivo study, proinflammatory lipopolysaccharide (LPS) and control disc nucleus surrogates (NP micromass pellets) were generated in vitro from rat NPCs and implanted into rat tail vertebrae, and the response was compared with sham surgery (n=12 each). Tissue changes were investigated with T1w and T2w MRI (7T), histology, and immunohistochemistry (tumor necrosis factor, CD3) 1 (n=6) and 2 weeks (n=6) after implantation. ResultsBMNC/NPC co-culture significantly increased lymphocyte viability (42%–69%, p<.05) and reduced NPC viability (96%–88%, p<.001), indicating immunogenicity of NPC. However, IL-1α was required to cause significant transcriptional upregulation of IL-1, IL-6, IL-10, and tropomyosin receptor kinase A. Therefore, an inflammatory activation is required to amplify the immune response. Immunogenicity of the NP was corroborated in vivo by CD3 cell accumulation around LPS and control disc surrogates at Day 7. However, only the LPS disc surrogate group demonstrated infiltration of CD3 cells at Day 14. Furthermore, end plate defects (p<.05, LPS: n=4/6, Ctrl: n=0/6, sham: n=0/6) and MC1-like MRI changes (T2w hyperintensity, p<.05) were only seen with LPS disc surrogates. ConclusionsNPCs are immunogenic but cannot trigger MC without an additional proinflammatory stimulus. Our data suggest that MC requires end plate defects that allow marrow/NPC co-mingling plus an adjacent inflammatory “MC disc” that can amplify the immune response.

Packed cell volume and bone marrow cytological responses in Trypanosoma vivax-induced acute trypanosomosis in Nigerian Sokoto Red goats

Bone marrow responses have been identified as key determinants of trypanotolerance in cattle as they determine the ability for hemopoietic cell regeneration and control of anemia. However, not much is known about such responses in Nigerian breed of goats known to show different susceptibilities to trypanosome infections. Early bone marrow events associated with pathogenesis of Sokoto Red goats to Trypanosoma vivax were investigated in six goats to assess their role in susceptibility of this goat breed to trypanosomiasis. A total of six Sokoto Red goats of mixed sexes were used. While four of the goats randomly selected were infected with Trypanosoma vivax, the remaining two served as control. The parameters examined included packed cell volume (PCV) and differential bone marrow cytology. Cytological changes in the bone marrow of goats with acute trypanosomosis were compared to the controls. T. vivax caused an acute disease course in the goats resulting to mild drop in PCV and death 2–3 weeks post infection (PI). Associated bone marrow (BM) cytological changes were characterized by moderate erythroid hyperplasia with a resultant lower myeloid:erythroid (M:E) ratio, while the granulocytic maturation rate was 3.15 ± 0.6 and 2.42 ± 0.0 for infected and control animals, respectively. However, significant macrophage (MC) numbers (hyperplasia) were detected in the BM of the infected group. Most of the MCs phagocytized mature red blood cells (RBC) and band or mature white blood cells (WBC) only while no MC phagocytized immature cells. The MCs in BM of control goats phagocytized no blood cells. The study confirmed that mature blood cells form the first cell types to be phagocytized by MCs of BM in the pathogenesis of anemia in T. vivax-infected Sokoto Red goats while early onset of MC hyperplasia and erythrophagocytosis are indicators of susceptibility to trypanosomosis in this breed of goats and perhaps other susceptible ruminants.

Femtosecond laser microstructured Alumina toughened Zirconia: A new strategy to improve osteogenic differentiation of hMSCs

The use of topographic patterns has been a continuously growing area of research for tissue engineering and it is widely accepted that the surface topography of biomaterials can influence and modulate the initial biological response. Ultrafast lasers are extremely powerful tools to machine and pattern the surface of a wide range of biomaterials, however, only few work has been performed on ceramics with the intent of biomedical applications, and the biological characterization of these structured materials is scarce.In this work, relevance is given to the biological performance of such materials. A femtosecond laser ablation technique was used to modify Alumina toughened Zirconia (ATZ) surface topography, developing surfaces structured at the micro and nanoscale levels (μATZ), in a controlled and reproducible manner. Materials characterization was performed before and after laser treatment, and both materials were compared in terms of osteogenic response of human bone marrow derived mesenchymal stem cells cultured under basal conditions, expecting that the micro/nanofeatures will improve the biological response of cells. Cells metabolic activity and proliferation increased with the culture time and surface microtopography modulated cells alignment and guided proliferation. The modified surface, displayed significantly higher expression of osteogenic transcription factors and genes and, additionally, the formation of a mineralized extracellular matrix, when compared to the control surface, i.e. unmodified ATZ.

Histomorphological responses after therapy with pegylated interferon \u03b1-2a in patients with essential thrombocythemia (ET) and polycythemia vera (PV)

BackgroundPegylated interferon alfa-2a (PEG-IFN-α-2a) is a potent immunomodulating agent capable of inducing high rate of hematologic and even complete molecular remission in patients with essential thrombocythemia (ET) and polycythemia vera (PV). We recently reported results of a phase 2 trial of PEG-IFN-α-2a in 83 patients with ET and PV after a median follow-up of 83 months. Here we report an analysis of bone marrow (BM) responses in these patients.MethodsAmong 83 patients, 58 (70%, PV 25, ET 31) had evaluable BM samples. BM responses and fibrosis grading were defined according to the International Working Group for Myeloproliferative Neoplasms Research and Treatment, and the European Consensus on grading of BM fibrosis, respectively. BM was assessed prior to enrollment, and every 6–24 months while on therapy in all patients, and after therapy discontinuation in some patients.ResultsThe median age of analyzed 58 patients was 52 years, and 29% were males. After a median follow-up of 84 months, 32 patients are still on study. Hematologic (HR) and molecular responses (MR) were seen in 93 and 69% patients, respectively. Twenty-nine patients (50%) had a BM response, including 13 (22%) with a complete BM response (BM-CR). Moreover, 13 patients (22%) have experienced complete resolution of bone marrow reticulin fibrosis. Patients with BM response had higher duration of HR and MR, and lower discontinuation rate. Furthermore, patients with BM-CR had a higher probability of complete MR. The median duration of BM-CR was 30 months, and 9 patients have maintained their BM-CR (69%), including five who have maintained their response after discontinuation of therapy. Despite this observation, the pattern of HR, MR and BM response, their durability and interrelation was heterogeneous.ConclusionsOur results show the ability of PEG-IFN-α-2a to induce complete BM responses in a subset of ET and PV patients, but its correlation with durable clinically relevant treatment benefit warrants further investigation.Trial registration This study is registered with ClinicalTrials.gov (NCT00452023), and is ongoing but not enrolling new patients.

The relationship between iron bone marrow stores and response to treatment in pediatric acute lymphoblastic leukemia

Iron is an intracellular element whose accumulation in the body is associated with tissue damage. This study examines the effect of iron on pediatric acute lymphoblastic leukemia (ALL) and its “response to treatment.” At the end of the first year of treatment, bone marrow iron store (BMIS) was evaluated in children with ALL and the relationship between iron store and minimal residual disease was investigated. Moreover, the 3-year disease-free survival (3-DFS) of patients was determined. Patients’ BMIS were compared with that of subjects with normal bone marrow. The study examined 93 children, including 78 Pre-B and 15 T-cell ALL patients. BMIS did not differ between the children with ALL and those with no evidence of cancer. BMIS was increased in 26.6% of patients at the end of the first year of treatment. Drug resistance and BM relapses were more prevalent in cases with high BMIS in both Pre-B and T-cell groups. Bone marrow iron store is not considered a risk factor for childhood ALL. However, high levels of BMIS are associated with poor response to treatment and the risk of relapse. Bone marrow iron store control during treatment can therefore help achieve better outcomes and improve the chances of recovery.

A regenerative erythropoietic response does not increase the frequency of Pig-a mutant reticulocytes and erythrocytes in Sprague-Dawley rats.

The in vivo rodent Pig-a mutation assay is a sensitive test to identify exposure to mutagenic substances, and has been proposed as an assay for the identification of impurities for pharmaceuticals. Red blood cells (RBCs) and reticulocytes (RETs) are analyzed by flow cytometry after exposure to potentially mutagenic chemicals for cells deficient in the cell surface anchored protein CD59, representing mutation in the X-linked Pig-a gene. The full potential of the assay as well as its limitations are currently being explored. The current study investigated the effects of regenerative erythropoietic bone marrow responses on the frequency of Pig-a mutated reticulocytes (RETCD59- ) and erythrocytes (RBCCD59- ). We hypothesized that a robust regenerative erythropoietic response would not increase the basal frequency of RETCD59- or RBCCD59- cells. Two groups of six male Sprague-Dawley rats either had 2 mL of blood sampled each day via an indwelling catheter over a period of 5 days or were minimally sampled for hematology and used as controls. Blood was also then collected and evaluated 5, 18, and 49 days after the initial bleed period for the number of Pig-a mutant cells in either the RET or RBC population. Despite the expected decrease in hematocrit and the correlative increase in reticulocytes after bleeding, no increase in the number of Pig-a mutant cells was observed in male Sprague-Dawley rats that were bled for five consecutive days. These results indicate that changes in erythropoiesis and hematology parameters in rats appear to have no effect on the background levels of Pig-a mutated RETs and RBCs. Environ. Mol. Mutagen. 59:91-95, 2018. © 2017 Wiley Periodicals, Inc.

Donor T-cell responses and disease progression patterns of multiple myeloma

Donor T-cells transferred after allogeneic stem cell transplantation (alloSCT) can result in long-term disease control in myeloma by the graft-versus-myeloma (GvM) effect. However, T-cell therapy may show differential effectiveness against bone marrow (BM) infiltration and focal myeloma lesions resulting in different control and progression patterns. Outcomes of 43 myeloma patients who underwent T-cell-depleted alloSCT with scheduled donor lymphocyte infusion (DLI) were analyzed with respect to diffuse BM infiltration and focal progression. For comparison, 12 patients for whom a donor search was started but no alloSCT was performed, were analyzed. After DLI, complete disappearance of myeloma cells in BM occurred in 86% of evaluable patients. The probabilities of BM progression-free survival (PFS) at 2 years after start of donor search, alloSCT and DLI, were 17% (95% confidence interval 0-38%), 51% (36-66%), and 62% (44-80%) respectively. In contrast, the probabilities of focal PFS at 2 years after start of donor search, alloSCT and DLI, were 17% (0-38%), 30% (17-44%) and 28% (11-44%), respectively. Donor-derived T-cell responses effectively reduce BM infiltration, but not focal progression in myeloma, illustrating potent immunological responses in BM with only limited effect of T-cells on focal lesions.

Balancing anti-inflammatory and anti-oxidant responses in murine bone marrow derived macrophages.

RationaleThe underlying pathophysiology of bronchopulmonary dysplasia includes a macrophage-mediated host response orchestrated by anti-inflammatory peroxisome proliferator-activated receptor gamma (PPARγ) and anti-oxidant nuclear factor (erythroid-derived 2)-like 2 (Nrf2). These have not yet been studied in combination. This study tested the hypothesis that combined inflammatory and oxidative stressors would interact and change PPARγ- and Nrf2-regulated gene expression and antioxidant capacity. Therefore, we investigated the effect of dual stimulation with lipopolysaccharide and hyperoxia in murine bone marrow-derived macrophages (BMDM).MethodsSub-confluent BMDM from wild-type C57BL/6J mice were treated with lipopolysaccharide (LPS) 1ug/mL for 2 hours followed by room air (21% oxygen) or hyperoxia (95% oxygen) for 24 hours. Taqman real time-polymerase chain reaction gene expression assays, total antioxidant capacity assays, and Luminex assays were performed.ResultsSupernatants of cultured BMDM contained significant antioxidant capacity. In room air, LPS treatment decreased expression of PPARγ and Nrf2, and increased expression of tumor necrosis factor-alpha and heme oxygenase-1; similar findings were observed under hyperoxic conditions. LPS treatment decreased cellular total antioxidant capacity in room air but not in hyperoxia. Increased expression of sulfiredoxin-1 in response to hyperoxia was not observed in LPS-treated cells. Dual stimulation with LPS treatment and exposure to hyperoxia did not have synergistic effects on gene expression. Cellular total antioxidant capacity was not changed by hyperoxia exposure.ConclusionsOur hypothesis was supported and we demonstrate an interaction between inflammatory and oxidative stressors in a model system of bronchopulmonary dysplasia pathogenesis. The protective anti-oxidant effect of cell culture media may have protected the cells from the most deleterious effects of hyperoxia.

The early response cells to bone marrow injury

We have previous studied the bone marrow response in submyeloablatively (6 Gy) irradiated mice. This allowed a detailed analysis of immature Lin-c-Kit+ (LK) cells only after approximately two weeks. These studies revealed significant expansion of Sca-1+ (S+) LK CD150+CD48+ cells with decreased c Kit expression level (Blood 2014 124:5112). Furthermore, they showed that not only LS+K cells responded to the injury but also Sca 1- (S-) late myeloid progenitors were activated and some of them were induced to re-expressed Sca 1 antigen. In present study, we focused on the response of LK cells occurring shortly after bone marrow damage by analyzing bone marrow 1-5 days after irradiation of mice with 2-4 Gy. The number of bone marrow cells significantly decreased after 2-4 Gy irradiation during one day. In LK cells, the LS-K/LS+K ratio decreased (below 2) due to Sca-1 induction. However, it recovered to normal level in a range of 5-7 after 3-5 days. We used CD150 and CD48 SLAM markers to distinguish between various subtypes of LSK cells, and CD34 and CD16/32 markers to characterize various types of LS-K myeloid progenitor cells. This revealed an early activation of megakaryocyte-erythroid and granulocyte macrophage myeloid progenitors. The activation was characterized by re-expression of Sca 1 in a part of these cells and by significantly decreased c-Kit expression level. During 3-5 days, c Kit recovered fully in 2 Gy- and partly in 4 Gy-irradiated mice. The transcription factor SCL/TAL1 with other co-factors is known to upregulate c-Kit gene and c-Kit expression. Therefore, we examined SCL/TAL1 mRNA level in LK cells in bone marrow of irradiated mice. Necas E, Szikszai Forgacova K, Faltusova K, et al: Lin-Sca-1+c-KitlowCD48+CD71+ Cells Are the Engine of Bone Marrow Regeneration. Blood 2014 124:5112 (abstract). Research was supported by the Grant Agency of the Czech Republic (GACR 17-01897S).

Transcriptomic biomarkers of altered erythropoiesis to detect autologous blood transfusion.

Autologous blood transfusion is a powerful means of improving performance and remains one of the most challenging methods to detect. Recent investigations have identified 3 candidate reticulocytes genes whose expression was significantly influenced by blood transfusion. Using quantitative reverse transcription polymerase chain reaction as an alternative quantitative method, the present study supports that delta-aminolevulinate synthase 2 (ALAS2), carbonic anhydrase (CA1), and solute carrier family 4 member 1 (SLC4A1) genes are down-regulated post-transfusion. The expression of these genes exhibited stronger correlation with immature reticulocyte fraction than with reticulocytes percentage. Moreover, the repression of reticulocytes' gene expression was more pronounced than the diminution of immature reticulocyte fraction and reticulocyte percentage following blood transfusion. It suggests that the 3 candidate genes are reliable predictors of bone marrow's response to blood transfusion and that they represent potential biomarkers for the detection of this method prohibited in sports.