Bone Marrow Proliferation Research Articles

Myasthenia Gravis and Myeloproliferative Neoplasms - Mere Association or Paraneoplastic Neurologic Syndrome: A Mini-Review.

Myasthenia Gravis (MG) is a rare neurological condition characterized by muscle weakness that worsens after use. Myeloproliferative Neoplasms (MPNs) are disorders due to stem-cell hyperplasia characterized by an increased peripheral blood cell count, overactive bone marrow, and proliferation of mature hematopoietic cells. MPNs may be Philadelphia (Ph) chromosome-positive or Negative. A systematic review of case reports was conducted by searching PubMed, Scopus, and Google scholar to identify case reports in which there is an association between MG and MPN and know whether MG can be considered a possible neurological paraneoplastic syndrome in patients with MPNs. A total of 13 cases of MPNs associated with MG were identified. The most common type of MPN associated with MG was chronic myeloid leukemia (CML) (10 out of 13 patients). In most of the patients, MG symptoms appeared after a diagnosis of MPN was made. Considering that 10 out of the 13 patients in our cohort had positive auto-antibodies though only 4 of them had thymic hyperplasia, we hypothesize that bone marrow proliferation was responsible for the production of autoantibodies in these patients.As the clonal cell population cannot be eliminated entirely in the bone marrow even after treatment with tyrosine kinase inhibitors (TKI) in Ph +ve MPNs and JAK2 inhibitors in Ph -ve MPNS, MG can occur even in patients who are treated with these agents. A high index of suspicion is needed to diagnose it early, and treatment should be initiated immediately with steroids and anticholinergic agents. (www.actabiomedica.it)

Proteomic analysis of the bone marrow microenvironment in extramedullary multiple myeloma patients.

Multiple myeloma (MM) is a heterogeneous hematological malignancy characterized by the uncontrolled clonal proliferation of bone marrow (BM) plasma cells. The poor prognosis of patients is associated with the presence of extramedullary disease (EMD). Previously, different mechanisms involved in the colonization of BM niches by MM cells and their escape during EMD have been described. Thus, we aimed to investigate the expression of selected cytokines in the BM plasma of MM patients as well as EMD patients to reveal novel molecules involved in EMD pathogenesis. Expression of 120 different cytokines was measured in BM plasma of 13 MM and 11 EMD patients using Proteome Profiler Antibody Arrays. The correlation between statistically significant cytokines and clinicopathological parameters of patients was determined using the Spearman correlation analysis. Finally, protein-protein interactions were analyzed, and GO and KEGG pathways enrichment analysis was performed. In total, 27 cytokines were found to be differently expressed between MM and EMD patients. After the Benjamini-Hochberg correction for multiple testing, the statistical significance of two cytokines downregulated in EMD (EGF, BDNF) and six cytokines upregulated in EMD (NAP-2, ADIPOQ, CRP, MIG, BAFF, and THBS1) was maintained. Correlation analysis proved a significant association between the expression of these molecules and selected clinical-pathological features of MM/EMD patients. Protein association network analysis revealed important protein-protein interactions between THBS1/EGF, MIG/NAP-2, THBS1/NAP-2, EGF/NAP-2, and ADIPOQ/CRP. Finally, identified cytokines were proved to be significantly involved in focal adhesion, PI3K/AKT, and MAPK signaling pathways, and regulation of cell development, localization, proliferation, migration, differentiation, immune system processes, and stress response. Obtained results confirm the key function of the BM microenvironment in the pathogenesis of MM and indicate the essential role of numerous cytokines in disease progression and EMD development. However, the exact mechanisms need to be further clarified.

Antibacterial wollastonite supported excellent proliferation and osteogenic differentiation of human bone marrow derived mesenchymal stromal cells

Biocompatibility and bacterial infections are the primary concerns associated with the current bone graft substitutes. The application of wollastonite-based scaffolds for bone tissue engineering becomes a novel subject of interest. In the present study, a single phasic wollastonite scaffold was synthesised using citric acid-based sol–gel combustion route. Its physicochemical characteristics, antibacterial properties as well as its biocompatibility and osteogenic induction effect on human bone marrow derived stromal cells (hBMSCs) are yet to be explored. The TGA/DTA, XRD and SEM/EDX confirmed the characteristics of wollastonite. The antibacterial test indicated wollastonite inhibition of 47.81% and 45.54% for gram-positive, Staphylococcus aureus and Staphylococcus epidermidis and 47.04% and 46.07% for gram-negative, Escherichia coli and Pseudomonas aeruginosa bacterial strains, respectively. The SEM micrographs demonstrated an excellent attachment of hBMSCs on wollastonite and comparable to commercial hydroxyapatite (cHA) scaffold. The alamar blue cell proliferation assay confirmed 1.7- and 1.8-fold significant increase in hBMSCs seeded on wollastonite and cHA scaffold, respectively, on day 14 as compared with day 1. The immunohistochemistry analysis on Type-I collagen (Col1) and Bone morphogenetic protein-2 (BMP2) expression on day 14 confirmed the osteogenic differentiation of hBMSCs seeded on wollastonite and comparable with cHA scaffold. In conclusion, wollastonite scaffold has a greater potential to substitute bone grafts in orthopaedic applications.

Abstract 14015: Extracellular Vesicle-Associated microRNAs From the Plasma of HIV-Positive Individuals Enhance Atherosclerosis Burden

Introduction: People living with HIV (PLHIV) have a higher risk of cardiovascular diseases (CVD) compared to HIV negative (HIV - ) individuals. The molecular mechanisms underlying the increased CVD risk remain elusive. Extracellular vesicles (EVs) mediate cell-to-cell communication through cargo transfer, including microRNAs (EV-miRs). Hypothesis: HIV infection results in modification of plasma EVs cargo and these HIV positive EVs (HIV posEVs ), specifically EV-miRs, recapitulate the effects of HIV infection on CVD development. Methods: Atherosclerosis burden was assessed by carotid ultrasound in 74 PLHIV treated with antiretroviral therapy (ART) with viral loads <50 c/ml and 23 HIV - subjects. Circulating endothelial progenitor cell (EPC) levels were measured by colony forming unit (CFU) assay. PLHIV plasma was fractionated into plasma EVs (HIV posEVs ) and plasma-depleted of EVs (HIV plasmadepEVs ). These fractions, EVs from HIV - subjects (HIV negEVs ) and PBS were injected weekly into apoE -/- mice via tail vein for three months. Atherosclerosis burden (H&E and Oil Red O staining), bone marrow (BM) EPC senescence (β-galactosidase staining), proliferation (MTT assay) and migration (“scratch” assay) were examined. RNASeq was used to identify EV-miRs that mediated the effects of HIV posEVs . Tailored EVs (TEVs) loaded with antagomirs for top EV-miRs were generated to determine their capacity to reverse the adverse effects of HIV posEVs in mouse EPCs in vitro . Results: PLHIV had higher atherosclerosis burden and lower circulating EPC numbers. Mice treated with HIV posEVs showed increased atherosclerotic burden, higher BM EPC senescence and lower functionality. HIV posEVs also negatively affected mouse BM EPC senescence in vitro . RNAseq identified four top candidate EV-miRs upregulated in HIV posEVs : let-7b-5p, miR-16-5p, miR-423-5p, and miR-103a-2. TEVs loaded with the antagomirs for let-7b and miR103a-2 counteracted the effects of HIV posEVs on the accelerated senescence of mouse BM EPCs. Conclusion: 1) Plasma EVs from PLHIV reconstitute the effects of HIV infection on accelerated atherosclerosis via negatively affects EPC functionality; and 2) TEVs, targeting candidate EV-miRs counteracts the effects of HIV infection on EPCs.

Bone Marrow-Based Biomarkers for Predicting aGVHD Using Targeted RNA Next Generation Sequencing and Machine Learning

Bone Marrow-Based Biomarkers for Predicting aGVHD Using Targeted RNA Next Generation Sequencing and Machine Learning

Alrn-6924, a Dual Inhibitor of MDMX and MDM2, Transiently Induces Cell Cycle Arrest in Bone Marrow and Prevents Toxicity in Mouse Models of Acute Radiation Injury

Background: Myelosuppression is a common sequela of acute radiation injury due to sensitivity of proliferating bone marrow cells to ionizing radiation. ALRN-6924 is a clinical-stage, first-in-class, stabilized cell-permeating alpha-helical peptide drug that disrupts the interaction of the p53 tumor suppressor protein with its endogenous inhibitors, MDMX and MDM2, to induce transient, dose-dependent cell cycle arrest in p53-wild-type tissues. ALRN-6924 is being evaluated in clinical trials as a selective chemoprotection agent for patients with p53-mutant cancers to protect healthy normal cells from chemotherapy while not protecting p53-mutant cancer cells. We tested whether ALRN-6924 may similarly protect against radiation-induced toxicity in mouse models of acute radiation injury.Materials and methods: Activation of p21 (CDKN1A), a cell cycle regulator under transcriptional control of p53, was measured in formalin-fixed mouse bone marrow by immunohistochemistry analysis (IHC). Proliferation and apoptosis in bone marrow were measured by IHC of Ki67 and cleaved PARP, respectively. Cell cycle arrest was measured in the bone marrow of ALRN-6924-treated C57BL/6 mice by flow cytometry using EdU incorporation. Serum levels of macrophage inhibitory cytokine-1 (MIC-1), a biomarker of p53 activation, were measured by ELISA. As a model of radiation-induced toxicity, C57BL/6 mice (n=7/group) were treated with one or more intravenous 2.4 mg/kg doses of ALRN-6924 at 24, 16, 8, or 1 hour (or combinations thereof) or placebo prior to a 15 Gy shielded-body radiation dose and then monitored for body weight (BW) changes.Results: ALRN-6924 induced cell cycle arrest in mouse bone marrow with a maximum effect at 8 hrs post-dose. Repeated doses of ALRN-6924 every 8 hrs elevated p21 levels in bone marrow that correlated with reduced Ki67 positivity and increased serum MIC-1 levels. Treatment-dependent changes in cPARP expression in bone marrow were evident, but minimal in magnitude. In a nonlethal radiation exposure model, ALRN-6924 yielded significant protection from radiation-induced BW loss in a schedule-dependent manner. Placebo-treated mice showed 10% to 15% BW loss five days after irradiation, while mice receiving one or more ALRN-6924 doses 8 hrs prior to irradiation had an average of 4% BW loss (p=0.008, two-sided t test).Conclusions: ALRN-6924 mitigated toxicity in a mouse model of acute radiation injury. The observed radioprotection effect was correlated with cell cycle arrest in bone marrow after one or more doses of ALRN-6924. These results support further investigation of ALRN-6924 as a radioprotective agent. DisclosuresAnnis: Aileron Therapeutics, Inc.: Current Employment. Sutton: Aileron Therapeutics, Inc.: Consultancy; Kriya Therapeutics: Consultancy; First Light Pharmaceuticals: Consultancy; Cygnal Therapeutics: Consultancy. Aivado: Aileron Therapeutics, Inc.: Current Employment. Vukovic: Aileron Therapeutics, Inc.: Current Employment.

Spatial Transcriptomics Reveals DPP4 As Novel Marker of a More Proliferative Phenotype in Early AML Progression

Acute myeloid leukemia (AML) is a hematologic malignancy with poor prognosis for which the standard-of-care chemotherapy treatment regimen has remained virtually unchanged over the past 40 years. We have employed “Image-Seq”, a new technology that was developed in our laboratory, to study spatial variations in early leukemia progression in a mouse model of HoxA9-Meis1 AML. We visualized leukemia cells with differing proliferative phenotype using intravital microscopy, captured these cells under image guidance from individual bone marrow microenvironments and studied their differential expression by single-cell RNA sequencing. This analysis identified DPP4 as a key upregulated gene in AML cells from more proliferative bone marrow compartments and associated DPP4 expression with a cell cluster enriched in progenitor cell markers for HoxA9-Meis1 AML, including Flt3, Itgb7 and Ddx4. Strikingly, DPP4 is not expressed in vitro, and its expression in vivo (as quantitated by FACS analysis) correlated with disease progression and marked a more proliferative phenotype both at the 1-week and 2-week time-points during disease progression. DisclosuresSykes: Clear Creek Bio: Current equity holder in publicly-traded company; SAFI Biosolutions: Consultancy, Current equity holder in publicly-traded company; Keros Therapeutics: Consultancy. Scadden: Magenta Therapeutics: Current holder of individual stocks in a privately-held company, Membership on an entity's Board of Directors or advisory committees; VCanBio: Consultancy; LifeVaultBio: Current holder of individual stocks in a privately-held company, Membership on an entity's Board of Directors or advisory committees; Inzen Therapeutics: Membership on an entity's Board of Directors or advisory committees; Garuda Therapeutics: Current holder of individual stocks in a privately-held company, Membership on an entity's Board of Directors or advisory committees; FOG Pharma: Consultancy; Fate Therapeutics: Current holder of individual stocks in a privately-held company; Editas Medicines: Current holder of individual stocks in a privately-held company, Membership on an entity's Board of Directors or advisory committees; Dainippon Sumitomo Pharma: Other: sponsored research; Clear Creek Bio: Current holder of individual stocks in a privately-held company, Membership on an entity's Board of Directors or advisory committees; Agios Pharmaceuticals: Current holder of individual stocks in a privately-held company, Membership on an entity's Board of Directors or advisory committees.

High Levels of Circulating Tumor Cells Are Associated with Increased Bone Marrow Proliferation in Newly Diagnosed Multiple Myeloma Patients

IntroductionThe introduction of new treatment regimens has significantly increased the progression free survival (PFS) of newly diagnosed multiple myeloma (MM) patients. However, even with these novel treatments, for some the disease remains refractory, highlighting the need to identify the pathobiology of high-risk MM. In MM patients, high levels of circulating tumor cells (CTCs) is associated with an inferior prognosis independent of high-risk cytogenetics (Chakraborty et al., 2016), suggesting that CTC numbers are a relevant reflection of tumor cell biology. We hypothesized that high levels of CTCs in MM patients are either the result of a transcriptionally distinct tumor clone with enhanced migration capacities, or driven by transcriptional differences present in the bone marrow (BM) tumor cells. To test these hypotheses, we 1) compared MM cells from paired blood and BM samples, and 2) compared BM tumor cells of patients with high and low CTC levels, using single cell RNA-sequencing.ResultsWe isolated plasma cell (PCs) from viably frozen mononuclear cells of paired peripheral blood (PB) and BM aspirates from five newly diagnosed MM patients (0.5%-8% CTCs) to determine the presence of a distinct CTC subclone. We generated single cell transcriptomes from 44,779 CTCs and 35,697 BM PCs. In the total 9 clusters common to BM PCs and CTCs were identified upon single cell data integration, but no cluster specific for either source was detected. Only 25 genes were significantly differential expressed between CTCs and BM PCs. The absence of transcriptional clusters unique to either CTCs or BM PCs, and the transcriptional similarity between these two anatomical sites makes it highly unlikely that CTC levels are driven by the presence of a transcriptionally-primed migratory clone.We next set out to identify possible transcriptional differences in BM PCs from eight patients with high (2-22%) versus thirteen patients with low (0.004%-0.08%) percentages of CTCs. Recurrent high-risk mutations were present in both groups. Single cell transcriptomes were generated from 74,830 BM PCs. Single cell data integration across all patients led to the identification of 8 distinct PC clusters, one of which was characterized by enhanced proliferation as defined by STMN1 and MKI67 transcription. Interestingly, this proliferative cluster was increased in patients with a high percentage of CTCs. Furthermore, cell cycle analyses based on canonical G2M and S phase markers revealed that actively cycling PCs were more frequent in the BM of patients with a high percentage of CTCs (64% versus 30%, p<0.001), irrespective of the transcriptional cluster of origin. We hypothesized that plasma cell-extrinsic cues from the bone marrow micro-environment might be driving tumor proliferation.In order to substantiate this, we isolated BM immune cells from the same 21 patients and generated a library of 301,045 single immune cell transcriptomes. This library contained all major immune cell subsets, including CD4 + and CD8 + T cells, NK cells, B cells and monocytes. Comparative analyses of these cell populations in patients with either high or low levels of CTC are ongoing.ConclusionThrough single cell transcriptomic analyses, we demonstrate that CTCs and BM PCs are transcriptionally similar. Importantly, we identify increased BM PC proliferation as a significant difference between patients with high and low levels of CTCs, implicating an increased tumor proliferation as one of the potential mechanisms driving CTC levels and MM disease pathobiology. The relation of the BM immune micro-environment to this altered proliferative state is currently under investigation. Disclosuresvan der Velden: Janssen: Other: Service Level Agreement; BD Biosciences: Other: Service Level Agreement; Navigate: Other: Service Level Agreement; Agilent: Research Funding; EuroFlow: Other: Service Level Agreement, Patents & Royalties: for network, not personally. Sonneveld: SkylineDx: Honoraria, Research Funding; Karyopharm: Consultancy, Honoraria, Research Funding; Amgen: Consultancy, Honoraria, Research Funding; Celgene/BMS: Consultancy, Honoraria, Research Funding; Janssen: Consultancy, Honoraria, Research Funding; Takeda: Consultancy, Honoraria, Research Funding. Broyl: Sanofi: Honoraria; Janssen Pharmaceuticals: Honoraria; Celgene: Honoraria; Bristol-Meyer Squibb: Honoraria; Amgen: Honoraria.

Murine Oncostatin M Has Opposing Effects on the Proliferation of OP9 Bone Marrow Stromal Cells and NIH/3T3 Fibroblasts Signaling through the OSMR.

The IL-6 family cytokine Oncostatin M (OSM) is involved in cell development, growth, hematopoiesis, inflammation, and cancer. Intriguingly, OSM has proliferative and antiproliferative effects depending on the target cell. The molecular mechanisms underlying these opposing effects are not fully understood. Previously, we found OSM upregulation in different myeloproliferative syndromes. However, OSM receptor (OSMR) expression was detected on stromal cells but not the malignant cells themselves. In the present study, we, therefore, investigated the effect of murine OSM (mOSM) on proliferation in stromal and fibroblast cell lines. We found that mOSM impairs the proliferation of bone marrow (BM) stromal cells, whereas fibroblasts responded to mOSM with increased proliferation. When we set out to reveal the mechanisms underlying these opposing effects, we detected increased expression of the OSM receptors OSMR and LIFR in stromal cells. Interestingly, Osmr knockdown and Lifr overexpression attenuated the OSM-mediated effect on proliferation in both cell lines indicating that mOSM affected the proliferation signaling mainly through the OSMR. Furthermore, mOSM induced activation of the JAK-STAT, PI3K-AKT, and MAPK-ERK pathways in OP9 and NIH/3T3 cells with differences in total protein levels between the two cell lines. Our findings offer new insights into the regulation of proliferation by mOSM.

Emerging roles of circular RNAs in osteoporosis.

Osteoporosis is one bone disease characterized with skeletal impairment, bone strength reduced and fracture risk enhanced. The regulation processes of bone metabolism are associated with several factors such as mechanical stimulation, epigenetic regulation and hormones. However, the mechanism of osteoporosis remains unsatisfactory. Increasing high‐throughput RNA sequencing and circular RNAs (circRNAs) microarray studies indicated that circRNAs are differentially expressed in osteoporosis. Growing functional studies further pinpointed specific deregulated expressed circRNAs (e.g., circ_28313, circ_0016624, circ_0006393, circ_0076906 and circ_0048211) for their functions involved in bone metabolism, including bone marrow stromal cells (BMSCs) differentiation, proliferation and apoptosis. Moreover, CircRNAs (circ_0002060, Circ_0001275 and Circ_0001445) may be acted as diagnostic biomarkers for osteoporosis. This review discussed recent progresses in the circRNAs expression profiling analyses and their potential functions in regulating BMSCs differentiation, proliferation and apoptosis.

Bone screws of porous silicon carbide coated with tantalum improve osseointegration and osteogenesis in goat femoral neck fractures

Traditional metal materials, such as stainless steel and titanium (Ti) alloys, are still the gold standards for fracture fixation. However, the elastic moduli of these materials differ from that of human cortical bone, and the stress shielding effect affects fracture healing, leading to secondary fractures. Herein, a new porous Ta coated SiC (pTa-SiC) scaffold using in internal fixation devices with good mechanical and biological properties was prepared based on porous silicon carbide (SiC) scaffold and tantalum (Ta) metal. The osteogenic and osseointegration properties of the pTa-SiC scaffold were investigated by both in vitro and in vivo tests. The results showed that compared with porous titanium (pTi), the pTa-SiC promoted the proliferation, migration, and osteogenic differentiation of human bone marrow–derived mesenchymal stem cells. Moreover, the internal fixation tests were carried out in a goat load-bearing femoral neck fracture model. Histological results showed good osseointegration around the pTa-SiC screws. And the acid etching results showed that bone cells grew tightly on the pTa-SiC throughout bone canaliculi, and the growth mode was contact osteogenesis, which indicated good biological fixation effects. Therefore, it is reasonable to be expected that the new pTa-SiC scaffold with excellent mechanical and biological properties could be a promising candidate for bone implant field.

Over-Expression of the Local Bone Marrow Renin-Angiotensin System in Acute Myeloid Leukemia.

Local bone marrow (BM) renin-angiotensin system (RAS) is an autocrine-paracrine system affecting hematopoiesis [Haznedaroglu IC, et al. A local RAS in the bone marrow. MedHypotheses. 1996; 46:507–10., Haznedaroglu IC. A local RAS in the bone marrow still awaits its Christopher Columbus. ExpHematol. 1998; 26:279., Haznedaroglu IC, et al.Towards the understanding of the local hematopoietic bone marrow RAS. IntJBiochemCellBiol. 2003; 35:867–80.] Angiotensin II type 1a (AT1a) receptors are present on the CD34+ hematopoietic stem cells from the BM or cord blood (CB) [Goker H, et al. Local umbilical cord blood renin-angiotensin system. AnnHematol . 2005; 84:277–81]. Angiotensin II stimulates the proliferation of BM and umbilical CB hematopoietic progenitors. Local RAS may also be involved in leukemogenesis and polycythemia vera [Aksu S, et. al. Enhanced Expression of the Local Haematopoietic Bone Marrow Renin-Angiotensin System in Polycythemia Rubra Vera J Int Med Res (in press), Aksu S, et. al. Over-expression of angiotensin converting enzyme (CD143) on leukemic blasts as a clue for the activated local bone marrow RAS in AML. Leukemia and Lymphoma (in press)]. Angiotensin converting enzyme (ACE) hyperfunction may lead to the acceleration of negative hematopoietic regulator peptide, AcSDKP, metabolism, which in turn lowers its level in the bone marrow microenvironment, finally removing the anti-proliferative effect of AcSDKP on the hematopoietic cells and blasts. Renin expression could have a role in leukemia development and angiotensin may act as an autocrine growth factor for AML cells. In this study, the expression of the mRNAs of the major RAS components, namely ACE, renin, and angiotensinogen in human BM samples were quantified by RT-PCR to confirm the presence of the local BM RAS. Furthermore, we assessed quantitative alterations of the ACE, renin, and angiotensinogen mRNAs in leukemic BM samples. The aim of this study was to search those major RAS components in the leukemic blast cells taken from the BM of the patients with AML. BM aspirations were obtained from 10 patients with AML and 8 patients with non-malignant hematological disorders. All of three gene expressions were found to be significantly higher in the BM samples of AML patients compared to the BM samples of the control group (Figure 1). Pathological activity of the local RAS-mediated regulation may be important, since the angiotensin peptides represent a molecular target in the disease management. [Display omitted]

Acrylic acid plasma polymerization and post-plasma ethylene diamine grafting for enhanced bone marrow mesenchymal stem cell behaviour on polycaprolactone nanofibers

Electrospun polycaprolactone (PCL) nanofibers (NFs) have been extensively researched for tissue engineering purposes, despite their hydrophobic surface properties which compromise their cell interactivity. To enhance this interactivity, within this study, the surface of PCL NFs was first modified by acrylic acid plasma polymerization. Scanning electron microscopy (SEM) and atomic force microscopy (AFM) confirmed that the plasma process did not damage the NFs. Water contact angle (WCA) and zeta potential measurements showed a significant wettability increase and surface charge decrease, respectively. X-ray photoelectron spectroscopy (XPS) confirmed that the NFs were coated with a plasma polymer containing O-based functional groups, which could be related to the aforementioned changes in surface wettability and charge. Additionally, a post-plasma grafting step of ethylene diamine on the surface carboxylic acid groups was performed to investigate the influence of COOH-groups on bone marrow mesenchymal stem cell (BMST) adhesion and proliferation. WCA, zeta potential and XPS measurements confirmed the successful conversion of carboxylic acid into primary amine groups, while SEM indicated that the NF morphology did not change upon the reaction. PCL NFs subjected to this post-plasma grafting evoked the best BMST behaviour, indicating that minor changes in the surface chemistry can have a significant effect on cell-biomaterial interactions.

Haemopoietic Potential of Tannin Fractionates of Vitex doniana Leaf on Nitrosobis (2-Oxopropyl) Amine Comobidity in Docetaxel-Induced Myelosuppression in Wistar Rat

Aim: This study investigated the haemopoietic potential of tannin fractionate of Vitex doniana leaf against nitosobis (2-oxopropyl) amine comorbidity in docetaxel-induced bone marrow suppression.&#x0D; Study Design: This is an experimental research.&#x0D; Place of Research: University of Nigeria, Enugu campus.&#x0D; Methodology: The male Wistar rats used in this experiment were twenty-eight in number, and they were grouped into 7, with each group having four rats. Group 1 served as control, and received 1ml of normal saline, while groups 2-7 were treated with Nitrosobis (2-oxopropyl) amine 5 mg/kg daily for 2 weeks. Then groups 3-7 were treated with 8 mg/Kg of docetaxel weekly for 2 weeks. And groups 4, 5 and 6 also received 250 mg/Kg and 500 mg/Kg and 1000 mg/kg of tannin, respectively, daily for 2 weeks. Group 7 received 40 mg/Kg of fesolate daily for 2 weeks.&#x0D; Results: The haemoglobin concentration and white blood cell count of rats in the groups treated with Nitrosobis (2-oxopropyl) amine alone (group 2) and Nitrosobis (2-oxopropyl) amine plus Docetaxel (group 3) showed statistically significant reduction (p=.05) in number when compared with the group treated with normal saline (group 1). The haemoglobin concentration and white blood cell count of the rats in the groups treated with 250 mg/kg, 500 mg/kg, and 1000 mg/kg of the tannin fractionate, in addition to the Nitrosobis (2-oxopropyl) amine and Docetxel (i.e. groups 4, 5, and 6, respectively) showed statistically significant dose dependent increase in number, with the group treated with 1000mg/kg showing the highest increment (p=.05). The cells in the bone marrow show significant reduction in number in the rats treated with Nitrosobis (2-oxopropyl) amine (group 2) and Nitrosobis (2-oxopropyl) amine plus docetaxel (group 3) when compared with the rats in group 1 (treated with normal saline). With the addition of graded doses of the tannin fractionate, 250 mg/kg, 500 mg/kg, and 1000mg/kg (i.e. groups 4, 5, and 6, respectively), the number of cells in the bone marrow showed statistically significant increase when compared to group 3 (treated with Nitrosobis (2-oxopropyl) amine plus docetxel), with the rats in the group treated with 1000 mg/kg of tannin fractionate (group 6) having the highest increment (p=.05).&#x0D; Conclusion: Tannin obtained from Vitex doniana leaf extract increases the haemoglobin concentration in dose dependent manner. It also increases the white blood cell count, and number of proliferating bone marrow cells, following suppression by Nitrosobis (2-oxopropyl) amine and Docetaxel. So, this tannin obtained from Vitex doniana leaf extract may be useful in clinical practice to cushion the myelosupression, anaemia and leukopenia that are associated with use of docetaxel in treatment of malignancies.

The Effect of Artemisia Dracunculus L. on Mitotic Index in Bone Marrow and Spleen Cells of Mice: In Vivo Study

Artemisia dracunculus L. (tarragon) is a rich source of herbal remedies with antioxidant and anti-inflammatoryproperties. In the present study, the proliferation of bone marrow and spleen cells of mice was evaluated afterextraction of Artemisia dracunculus with 80% of ethanol. Two doses of the extract (500mg/kg body weightand 1000mg/kg body weight) were given to the mice for seven successive days. On day eight, mice weresacrificed and cells from bone marrow and spleen were collected; mitotic index was calculated and resultswere compared with that of methotrexate at a dose 20mg/kg body weight ( positive control ) and distilledwater ( negative control). The results showed that the dose 500mg/kg body weight of Artemisia dracunculusextract caused significant increase in mitotic index in both bone marrow and spleen cells of mice whencompared with negative control. While, the dose 1000mg/kg body weight of Artemisia dracunculus extractcaused significant decrease in mitotic index in both bone marrow and spleen cells of mice when comparedwith negative control.

Reactive oxygen species (ROS)-responsive biocompatible polyethylene glycol nanocomposite hydrogels with different graphene derivatives

Reactive oxygen species (ROS) are important in regulating normal cell physiological functions, i.e., cell death, proliferation and differentiation. Redox modulation could have significant implications providing an opportunity for the development of new strategies to improve clinical therapeutic outcomes in the treatment of diabetes, hypertension, atherosclerosis, carcinogenesis, aging and infections. Here, we report a versatile synthetic method to produce three polyethylene glycol (PEG)-based nanocomposite hydrogels containing different graphene derivatives homogeneously distributed: graphene oxide (GO), reduced graphene oxide (rGO) and graphene nanoplatelets (GNP) by free-radical redox polymerization. The graphene–PEG nanocomposite hydrogels (GCH) were very stable at different pH and solvents. Incorporation of small amount (1% wt) of graphene additives led to enhanced mechanical properties, up to 2.5-fold increase in elastic modulus and higher thermal stability (53–62 °C). The swelling behavior strongly depended on the functionalization of the graphene additive and their interaction with PEG. Interestingly, incorporation of graphene additives conferred antioxidant/pro-oxidant activity to the hydrogel, with their radical scavenging activity depending on the nature of the radical and the graphene derivative. PEG-rGO and PEG-GNP showed the highest scavenging activity for 2,2-Diphenyl-1-picrylhydrazyl radical (DPPH•) and hydroxyl radical, respectively. In addition, PEG-rGO demonstrated peroxidase activity in the presence of H2O2. The three GCH proved biocompatible, with no effect on cell viability and proliferation of human bone marrow derived mesenchymal stem cells (hMSC). The results pave the way for the design of bioactive functional nanocomposite hydrogels for ROS-mediated applications.

Trilaciclib dose selection: an integrated pharmacokinetic and pharmacodynamic analysis of preclinical data and Phase Ib/IIa studies in patients with extensive-stage small cell lung cancer

PurposeTrilaciclib is a first-in-class CDK4/6 inhibitor that transiently arrests hematopoietic stem and progenitor cells (HSPCs) in the G1 phase of the cell cycle to preserve them from chemotherapy-induced damage (myelopreservation). We report integrated analyses of preclinical and clinical data that informed selection of the recommended Phase II dose (RP2D) used in trilaciclib trials in extensive-stage small cell lung cancer (ES-SCLC).MethodsA semi-mechanistic pharmacokinetic/pharmacodynamic (PK/PD) model developed from preclinical data guided selection of an optimal dose for G1 bone marrow arrest in a first-in-human Phase I study (G1T28-1-01). PK, PD, safety, and efficacy data from G1T28-1-01 and two Phase Ib/IIa studies (G1T28-02/-03) in ES-SCLC were analyzed to support RP2D selection.ResultsModel simulation of bone marrow arrest based on preclinical data predicted that a ≥ 192 mg/m2 dose would induce a 40–50% decrease in total bone marrow proliferation in humans and almost 100% cell cycle arrest of cycling HSPCs. Consistent with this model, analysis of bone marrow aspirates in healthy volunteers after trilaciclib 192 mg/m2 administration demonstrated almost 100% G1 arrest in HSPCs and 40% decrease in total bone marrow proliferation, with minimal toxicity. G1T28-02/-03 reported similar PK parameters with trilaciclib 200 mg/m2 but slightly lower exposures than expected compared with healthy volunteers; consequently, 240 and 280 mg/m2 doses were also tested to match healthy volunteer exposures. Based on PK and relevant safety data, 240 mg/m2 was selected as the RP2D, which was also favored by myelopreservation endpoints in G1T28-02/-03.ConclusionIntegrated PK/PD, safety, and efficacy data support 240 mg/m2 as the RP2D for trilaciclib.ClinicalTrials.gov IdentifiersNCT02243150; NCT02499770; NCT02514447.

Morphofunctional parameters of bone marrow of mice under using of antihypertensive agent

Due to the increase in cardiovascular disease, it is urgent to research new more effective and safe drugs and their combinations. Candesartan cilexetil is an angiotensin II receptor antagonist. It is known that candesartan cilexetil has a number of side effects, affecting the antioxidant system. Therefore, we studied the natural antioxidant resveratrol to prevent the cytotoxic effect. Goal. To study the effect of candesartan cilexetil and resveratrol in different doses on the number of apoptotic and micronucleated cells and cell cycle parameters in mice. Methods. Male C57Bl/6 mice were used for the experiment. Animals were received intragastrically candesartan cilexetil and resveratrol in different doses for 7 weeks. Evaluation of the effects of substances on cytogenetic and cytokinetic parameters of bone marrow was conducted by flow cytometry. Results. It was found that candesartan cilexetil at 1.5 mg/kg dose was cytotoxic. Using candesartan with resveratrol was safer for cells of bone marrow. Natural resveratrol reduced in combination the adverse effects of candesartan cilexetil. Test complex of substances stimulated cell proliferation in bone marrow of mice. Conclusions. New combination of candesartan cilexetil and resveratrol increased proliferation of cells and was not cytotoxic.

The multi-kinase inhibitor dasatinib suppresses autoinflammation and increases bone density in a mouse model for chronic recurrent multifocal osteomyelitis.

Chronic recurrent multifocal osteomyelitis (CRMO) is an autoinflammatory bone disease that presents with bone destruction and pain. Although genetic studies have identified signalling pathways involving CRMO, molecularly targeted drugs remain unavailable. We used an animal model of CRMO as an in vivo screening system for candidate therapeutic agents. A gain-of-function mutation in Fgr, a member of Src family kinases (SFKs), causes peripheral paw inflammation and reduced bone mineral density (BMD) in Ali18 mice. The SFK inhibitor dasatinib was selected for administration to Ali18 mice daily for 2 weeks. Local inflammation and BMD were assessed by clinical scoring and computed tomography, respectively. Pilot studies in a small number of animals showed that dasatinib administration effectively suppressed the early phase of autoinflammation in Ali18 mice. Serial oral gavage of dasatinib to a group of Ali18 mice confirmed significant suppression of paw swelling with no side effects. Histological analysis revealed that abnormal proliferative bone marrow cells and inflammatory infiltration into the skin in the affected area were clearly reduced in the animals with dasatinib administration. Further, trabecular BMD in Ali18 long bones was restored to levels similar to that found in wild type mice. Our results indicate that autoinflammation and related-bone phenotypes were completely suppressed by the dasatinib kinase inhibitor in CRMO model animals. Thus, it is strongly suggested that dasatinib can be used for clinical treatments of CRMO with the combination of molecular diagnosis of the FGR locus. SIGNIFICANCE OF THE STUDY: Autoinflammation and related-bone phenotypes were effectively suppressed by the kinase inhibitor dasatinib in CRMO model animals. In combination with molecular analysis of the FGR locus, dasatinib is a strong candidate for the clinical treatments of CRMO. We propose that the animal model employed in this study can be used to screen this and other potential drugs for CRMO.

LncRNA NORAD promotes bone marrow stem cell differentiation and proliferation by targeting miR-26a-5p in steroid-induced osteonecrosis of the femoral head

BackgroundSteroid-induced osteonecrosis of the femoral head (SONFH) is a devastating orthopedic disease, which seriously affects the quality of life of patients. The study aims to investigate the effects of LncRNA NORAD on SONFH.MethodsHuman bone marrow-derived mesenchymal stem cells (hBMSCs) were isolated from the proximal femur of patients during routine orthopedic surgery and then cultured with dexamethasone (Dex) and transfected with NORAD overexpression vector, siRNA-NORAD and miR-26a-5p mimics. The mRNA expression of NORAD, miR-26a-5p, OPG, RANK, and RANKL was detected by RT-qPCR. Cell proliferation and apoptosis was measured by CCK-8 assay and flow cytometry, respectively. The protein expression of RUNX2, OPG, RANK, and RANKL was detected by western blot. The dual-luciferase reporter gene assay was performed to confirm the binding between NORAD and miR-26a-5p.ResultsNORAD expression was downregulated in SONFH tissues, while miR-26a-5p expression was upregulated. Overexpression of NORAD improved DEX-induced inhibition of proliferation and differentiation, and promotion of apoptosis in hBMSCs, while knockdown of NORAD led to the opposite results. Moreover, NORAD improved DEX-induced inhibition of proliferation and differentiation, and promotion of apoptosis by regulation of miR-26a-5p in hBMSCs.ConclusionsNORAD expression was downregulated in SONFH tissues, while miR-26a-5p expression was upregulated. NORAD improved DEX-induced inhibition of proliferation and differentiation, and promotion of apoptosis by regulation of miR-26a-5p in hBMSCs.