Attenuation of cardiac hypertrophy by G-CSF is associated with enhanced migration of bone marrow-derived cells.

Granulocyte-colony stimulating factor (G-CSF) has been shown to promote mobilization of bone marrow-derived stem cells (BMCs) into the bloodstream associated with improved survival and cardiac function after myocardial infarction. Therefore, the aim of the present study was to investigate whether G-CSF is able to attenuate cardiac remodelling in a mouse model of pressure-induced LV hypertrophy focusing on mobilization and migration of BMCs. LV hypertrophy was induced by transverse aortic constriction (TAC) in C57BL/6J mice. Four weeks after TAC procedure. Mice were treated with G-CSF (100 µg/kg/day; Amgen Biologicals) for 2 weeks. The number of migrated BMCs in the heart was analysed by flow cytometry. mRNA expression and protein level of different growth factors in the myocardium were investigated by RT-PCR and ELISA. Functional analyses assessed by echocardiography and immunohistochemical analysis were performed 8 weeks after TAC procedure. G-CSF-treated animals revealed enhanced homing of VLA-4(+) and c-kit(+) BMCs associated with increased mRNA expression and protein level of the corresponding homing factors Vascular cell adhesion protein 1 and Stem cell factor in the hypertrophic myocardium. Functionally, G-CSF significantly preserved LV function after TAC procedure, which was associated with a significantly reduced area of fibrosis compared to control animals. Furthermore, G-CSF-treated animals revealed a significant improvement of survival after TAC procedure. In summary, G-CSF treatment preserves cardiac function and is able to diminish cardiac fibrosis after induction of LV hypertrophy associated with increased homing of VLA-4(+) and c-kit(+) BMCs and enhanced expression of their respective homing factors VCAM-1 and SCF.

Tumor necrosis factor stimulates osteoclastogenesis from human bone marrow cells under hypoxic conditions.

Bone homeostasis is maintained by the balance between osteoblastic bone formation and osteoclastic bone resorption. In this study, we used human bone marrow cells (BMCs) to investigate the role of hypoxic exposure on human osteoclast (OC) formation in the presence of tumor necrosis factor (TNF). Exposing the BMCs to 3%, 5%, or 10% O2 in the presence of receptor activator of NF-κB ligand (RANKL) and macrophage colony-stimulating factor (M-CSF) generated tartrate-resistant acid phosphatase (TRAP)-positive multinuclear cells, consistent with OCs. The addition of TNF under hypoxic conditions generated significantly greater numbers of mature OCs with more nuclei than OCs generated under normoxic conditions. Longer initial hypoxic exposure increased the number of OC precursor cells and facilitated the differentiation of OC precursor cells into multinucleated OCs. Quantitative RT-PCR analysis revealed that RANKL and TNFR1 were expressed at higher levels in non-OC cells from BMCs under hypoxic conditions than under normoxic conditions. Furthermore, to confirm the involvement of TNF-induced signaling, we examined the effects of blocking antibodies against TNFR1 and TNFR2 on OC formation under hypoxic conditions. The TNFR1 antibody was observed to significantly suppress OC formation. These results suggest that hypoxic exposure plays an important role in TNF-induced osteoclastogenesis from human BMCs.

Biological and Medicinal Chemistry Sector of The Royal Society of Chemistry: Promoting Chemistry Learning, Networking and Excellence for Baby Boomers through to Gen Alpha!

The Biological and Medicinal Chemistry Sector (BMCS) is an important interest group within the UK's Royal Society of Chemistry (RSC). Operating through a committee of voluntary members, the main goal of the BMCS is to share knowledge within the sector, primarily by organizing high quality scientific meetings, with a particular focus on networking. Financial support and tailored scientific programmes encourage training and development across multiple generations, from school age through to retirement. Scientific excellence is recognised through several high-profile awards.

Frequency of acute myeloid leukaemia-associated mouse chromosome 2 deletions in X-ray exposed immature haematopoietic progenitors and stem cells.

Exposure to ionising radiation can lead to an increased risk of cancer, particularly leukaemia. In radiation-induced acute myeloid leukaemia (rAML), a partial hemizygous deletion of mouse chromosome 2 is a common feature in several susceptible strains. The deletion is an early event detectable 24h after exposure in bone marrow cells using cytogenetic techniques. Expanding clones of bone marrow cells with chromosome 2 deletions can be detected less than a year after exposure to ionising radiation in around half of the irradiated mice. Ultimately, 15-25% of exposed animals develop AML. It is generally assumed that leukaemia originates in an early progenitor cell or haematopoietic stem cell, but it is unknown whether the original chromosome damage occurs at a similar frequency in committed progenitors and stem cells. In this study, we monitored the frequency of chromosome 2 deletions in immature bone marrow cells (Lin(-)) and haematopoietic stem cells/multipotent progenitor cells (LSK) by several techniques, fluorescent in situ hybridisation (FISH) and through use of a reporter gene model, flow cytometry and colony forming units in spleen (CFU-S) following ex vivo or in vivo exposure. We showed that partial chromosome 2 deletions are present in the LSK subpopulation, but cannot be detected in Lin(-) cells and CFU-S12 cells. Furthermore, we transplanted irradiated Lin(-) or LSK cells into host animals to determine whether specific irradiated cell populations acquire an increased proliferative advantage compared to unirradiated cells. Interestingly, the irradiated LSK subpopulation containing cells carrying chromosome 2 deletions does not appear to repopulate as well as the unirradiated population, suggesting that the chromosomal deletion does not provide an advantage for growth and in vivo repopulation, at least at early stages following occurrence.

Exploring the Potential Molecular Mechanisms of Interactions between a Probiotic Consortium and Its Coral Host.

Beneficial microorganisms for corals (BMCs) have been demonstrated to be effective probiotics to alleviate bleaching and mitigate coral mortality in vivo. The selection of putative BMCs is traditionally performed manually, using an array of biochemical and molecular tests for putative BMC traits. We present a comprehensive genetic survey of BMC traits using a genome-based framework for the identification of alternative mechanisms that can be used for future in silico selection of BMC strains. We identify exclusive BMC traits associated with specific strains and propose new BMC mechanisms, such as the synthesis of glycine betaine and ectoines. Our roadmap facilitates the selection of BMC strains while increasing the array of genetic targets that can be included in the selection of putative BMC strains to be tested as coral probiotics. IMPORTANCE Probiotics are currently the main hope as a potential medicine for corals, organisms that are considered the marine "canaries of the coal mine" and that are threatened with extinction. Our experiments have proved the concept that probiotics mitigate coral bleaching and can also prevent coral mortality. Here, we present a comprehensive genetic survey of probiotic traits using a genome-based framework. The main outcomes are a roadmap that facilitates the selection of coral probiotic strains while increasing the array of mechanisms that can be included in the selection of coral probiotics.

Coral Probiotics: Premise, Promise, Prospects.

The use of Beneficial Microorganisms for Corals (BMCs) has been proposed recently as a tool for the improvement of coral health, with knowledge in this research topic advancing rapidly. BMCs are defined as consortia of microorganisms that contribute to coral health through mechanisms that include (a) promoting coral nutrition and growth, (b) mitigating stress and impacts of toxic compounds, (c) deterring pathogens, and (d) benefiting early life-stage development. Here, we review the current proposed BMC approach and outline the studies that have proven its potential to increase coral resilience to stress. We revisit and expand the list of putative beneficial microorganisms associated with corals and their proposed mechanismsthat facilitate improved host performance. Further, we discuss the caveats and bottlenecks affecting the efficacy of BMCs and close by focusing on the next steps to facilitate application at larger scales that can improve outcomes for corals and reefs globally.

Evidence That the Adenovirus Single-Stranded DNA Binding Protein Mediates the Assembly of Biomolecular Condensates to Form Viral Replication Compartments.

A common viral replication strategy is characterized by the assembly of intracellular compartments that concentrate factors needed for viral replication and simultaneously conceal the viral genome from host-defense mechanisms. Recently, various membrane-less virus-induced compartments and cellular organelles have been shown to represent biomolecular condensates (BMCs) that assemble through liquid-liquid phase separation (LLPS). In the present work, we analyze biophysical properties of intranuclear replication compartments (RCs) induced during human adenovirus (HAdV) infection. The viral ssDNA-binding protein (DBP) is a major component of RCs that contains intrinsically disordered and low complexity proline-rich regions, features shared with proteins that drive phase transitions. Using fluorescence recovery after photobleaching (FRAP) and time-lapse studies in living HAdV-infected cells, we show that DBP-positive RCs display properties of liquid BMCs, which can fuse and divide, and eventually form an intranuclear mesh with less fluid-like features. Moreover, the transient expression of DBP recapitulates the assembly and liquid-like properties of RCs in HAdV-infected cells. These results are of relevance as they indicate that DBP may be a scaffold protein for the assembly of HAdV-RCs and should contribute to future studies on the role of BMCs in virus-host cell interactions.

Delivering Beneficial Microorganisms for Corals: Rotifers as Carriers of Probiotic Bacteria.

The use of Beneficial Microorganisms for Corals (BMCs) to increase the resistance of corals to environmental stress has proven to be effective in laboratory trials. Because direct inoculation of BMCs in larger tanks or in the field can be challenging, a delivery mechanism is needed for efficient transmission of the BMC consortium. Packaged delivery mechanisms have been successfully used to transmit probiotics to other organisms, including humans, lobsters, and fish. Here, we tested a method for utilizing rotifers of the species Brachionus plicatilis for delivery of BMCs to corals of the species Pocillopora damicornis. Epifluorescence microscopy combined with a live/dead cell staining assay was used to evaluate the viability of the BMCs and monitor their in vivo uptake by the rotifers. The rotifers efficiently ingested BMCs, which accumulated in the digestive system and on the body surface after 10 min of interaction. Scanning electron microscopy confirmed the adherence of BMCs to the rotifer surfaces. BMC-enriched rotifers were actively ingested by P. damicornis corals, indicating that this is a promising technique for administering coral probiotics in situ. Studies to track the delivery of probiotics through carriers such as B. plicatilis, and the provision or establishment of beneficial traits in corals are the next proof-of-concept research priorities.

miR-188 promotes senescence of lineage-negative bone marrow cells by targeting MAP3K3 expression.

Lineage-negative bone marrow cells (lin-BMCs) have reparative potential for overcoming endothelial dysfunction and reducing cardiovascular risk. Here, we found that miR-188 is upregulated and mitogen-activated protein kinase kinase kinase 3 (MAP3K3) is downregulated in aged lin-BMCs, whereas their expression is reversed in young lin-BMCs. We identified and confirmed MAP3K3 as a direct target of miR-188. MiR-188 overexpression or MAP3K3 silencing in young lin-BMCs increases p16 and p21 expression, enhances cell senescence, and decreases the ability for cell proliferation, migration, and tube formation. Conversely, miR-188 suppression in aged lin-BMCs yields the opposite results. We further found that MAP3K3 is involved in miR-188-induced promotion of lin-BMC senescence. All data reveal that miR-188 induces lin-BMC senescence by targeting MAP3K3 expression, thus, providing new theoretical basis for the prevention and treatment of cardiovascular diseases.

Effects of timing on intracoronary autologous bone marrow-derived cell transplantation in acute myocardial infarction: a meta-analysis of randomized controlled trials.

BACKGROUND: Several cell-based therapies for adjunctive treatment of acute myocardial infarction have been investigated in multiple clinical trials, but the timing of transplantation remains controversial. We conducted a meta-analysis of randomized controlled trials to investigate the effects of timing on bone marrow-derived cell (BMC) therapy in acute myocardial infarction (AMI). METHODS: A systematic literature search of PubMed, MEDLINE, and Cochrane Evidence-Based Medicine databases from January 2000 to June 2017 was performed on randomized controlled trials with at least a 3-month follow-up for patients with AMI undergoing emergency percutaneous coronary intervention (PCI) and receiving intracoronary BMC transfer thereafter. The defined end points were left ventricular (LV) ejection fraction, LV end-diastolic and end-systolic index. The data were analyzed to evaluate the effects of timing on BMC therapy. RESULTS: Thirty-four RCTs comprising a total of 2,307 patients were included; the results show that, compared to the control group, AMI patients who received BMC transplantation showed significantly improved cardiac function. BMC transplantation 3-7 days after PCI (+3.32%; 95% CI, 1.91 to 4.74; P < 0.00001) resulted in a significant increase of left ventricular ejection fraction (LVEF). As for the inhibitory effect on ventricular remodeling, BMC transplantation 3-7 days after PCI reduced LV end-diastolic indexes (-4.48; 95% CI, -7.98 to -0.98; P = 0.01) and LV end-systolic indexes (-6.73; 95% CI, -11.27 to -2.19; P = 0.004). However, in the groups who received BMC transplantation either within 24 hours or later than 7 days there was no significant effect on treatment outcome. In subgroup analysis, the group with LVEF ≤ 50% underwent a significant decrease in LV end-diastolic index after BMC transplantation (WMD = -3.29, 95% CI, -4.49 to -2.09; P < 0.00001); the decrease was even more remarkable in the LV end-systolic index after BMC transplantation in the group with LVEF ≤ 50% (WMD = -5.25, 95% CI, -9.30 to -1.20; P = 0.01), as well as in patients who received a dose of 10^7-10^8 cells (WMD = -12.99, 95% CI, -19.07 to -6.91; P < 0.0001). In the group with a follow-up of more than 12 months, this beneficial effect was significant and increased to a more pronounced effect of +3.58% (95% CI, 1.55 to 5.61; P = 0.0006) when compared with control. CONCLUSIONS: In this meta-analysis, BMC transfer at 3 to 7 days post-AMI was superior to transfer within 24 hours or more than 7 days after AMI in improving LVEF and decreasing LV end-systolic dimensions or LV end-diastolic dimensions. It is more effective in patients with lower baseline LVEF (≤50%) and the effect can last more than 12 months.

Cell Therapy for Liver Disease Using Bioimaging Rats.

Advances in stem cell research suggest that cell therapy is a potential alternative to liver transplantation. The use of individualized and minimally invasive cell therapy is desirable to avoid rejection and reduce patient burden. While allo-hepatocyte transplantation has been performed for metabolic hepatic disease, auto-bone marrow transplantation (BMT) has shifted toward mesenchymal stem cells (MSCs) transplantation for liver cirrhosis. In this article, an overview of cell transplantation research for liver disease is provided through our recent rat studies. We have developed various kinds of rat imaging models and have evaluated the effect of cell therapy for liver disease. Bone marrow cells (BMCs) of the Alb-DsRed2 rat were transplanted via the portal vein (PV) in acute and chronic liver damage models. The number of Alb-DsRed2+ albumin-producing cells increased, and the size of the cells increased in the chronic liver damage model as well as in the acute liver damage model. Luciferase transgenic (luc-Tg) rat hepatocytes were transplanted into the hepatectomized LEW rat via the PV. Luminescence intensity lasted for 2 months in the hepatectomized rat. BMCs obtained from green fluorescent protein (GFP) Tg rats were transplanted repeatedly via the PV using an implanted catheter with a port. Repeated BMT via the PV reduced the liver fibrosis. Adipocyte-derived MSCs from the luc-Tg rat were transplanted into the hepatectomized rat model via the PV after ischemic reperfusion. MSCs inhibited hepatocyte apoptosis and promoted liver regeneration. Transplanting the optimal number of cells by an effective and safe way is important for clinical application. Bioimaging rats are a powerful tool for cell transplantation research because it makes observation of the in vivo kinetics of transplanted cells possible. Cell transplantation research using bioimaging rats contributes greatly to evaluating effective methods of cell therapy.

Massive elimination of multinucleated osteoclasts by eupatilin is due to dual inhibition of transcription and cytoskeletal rearrangement.

Osteoporosis is an aging-associated disease requiring better therapeutic modality. Eupatilin is a major flavonoid from Artemisia plants such as Artemisia princeps and Artemisia argyi which has been reported to possess various beneficial biological effects including anti-inflammation, anti-tumor, anti-cancer, anti-allergy, and anti-oxidation activity. Complete blockade of RANK-dependent osteoclastogenesis was accomplished upon stimulation prior to the receptor activator of nuclear factor κB (RANK)-ligand (RANKL) treatment or post-stimulation of bone marrow macrophages (BMCs) in the presence of RANKL with eupatilin. This blockade was accompanied by inhibition of rapid phosphorylation of Akt, GSK3ß, ERK and IκB as well as downregulation of c-Fos and NFATc1 at protein, suggesting that transcriptional suppression is a key mechanism for anti-osteoclastogenesis. Transient reporter assays or gain of function assays confirmed that eupatilin was a potent transcriptional inhibitor in osteoclasts (OC). Surprisingly, when mature osteoclasts were cultured on bone scaffolds in the presence of eupatilin, bone resorption activity was also completely blocked by dismantling the actin rings, suggesting that another major acting site of eupatilin is cytoskeletal rearrangement. The eupatilin-treated mature osteoclasts revealed a shrunken cytoplasm and accumulation of multi-nuclei, eventually becoming fibroblast-like cells. No apoptosis occurred. Inhibition of phosphorylation of cofilin by eupatilin suggests that actin may play an important role in the morphological change of multinucleated cells (MNCs). Human OC similarly responded to eupatilin. However, eupatilin has no effects on osteoblast differentiation and shows cytotoxicity on osteoblast in the concentration of 50 µM. When eupatilin was administered to LPS-induced osteoporotic mice after manifestation of osteoporosis, it prevented bone loss. Ovariectomized (OVX) mice remarkably exhibited bone protection effects. Taken together, eupatilin is an effective versatile therapeutic intervention for osteoporosis via; 1) transcriptional suppression of c-Fos and NFATc1 of differentiating OC and 2) inhibition of actin rearrangement of pathogenic MNCs.

Delivery and processing of exogenous double-stranded DNA in mouse CD34+ hematopoietic progenitor cells and their cell cycle changes upon combined treatment with cyclophosphamide and double-stranded DNA.

We previously reported that fragments of exogenous double-stranded DNA can be internalized by mouse bone marrow cells without any transfection. Our present analysis shows that only 2% of bone marrow cells take up the fragments of extracellular exogenous DNA. Of these, ~45% of the cells correspond to CD34+ hematopoietic stem cells. Taking into account that CD34+ stem cells constituted 2.5% of the total cell population in the bone marrow samples analyzed, these data indicate that as much as 40% of CD34+ cells readily internalize fragments of extracellular exogenous DNA. This suggests that internalization of fragmented dsDNA is a general feature of poorly differentiated cells, in particular CD34+ bone marrow cells. When linearized plasmid DNA was used as a source of exogenous DNA, we observed that exonucleolytic processing and ligation of double-stranded DNA termini occurred in the bone marrow cells that had this DNA internalized. We also recovered "hybrid" plasmids that encompass kanamycin-resistance gene from the exogenous plasmid DNA and the fragments of plasmids from host enterobacteria, which is suggestive of recombination events taking place upon DNA internalization. CD34+ cells make up the distinctive bone marrow cell population that internalizes extracellular DNA. Cell cycle analysis of CD34+ cells treated with cyclophosphamide only or in combination with dsDNA, suggests that these cells have distinct biologic responses to these treatments. Namely, whereas upon cyclophosphamide treatment bone marrow stem cells become arrested at S-G2 phases, combined cyclophosphamide+dsDNA treatment leads to cell cycle progression without any delay. This indicates that when the genome is undergoing repair of interstrand crosslinks, injection of fragmented exogenous dsDNA results in immediate reconstitution of genome integrity. We observe that cyclophosphamide-only or a combined cyclophosphamide+dsDNA treatment of cells lead to two distinct waves of apoptosis in CD34+ progenitors. We also show that cyclophosphamide and cyclophosphamide+dsDNA injections promote division of CD34+ cells at distinct time periods.

Anti-thrombosis effect of LRRFIP1 shRNA lentivirus in a mouse model of deep vein thrombosis.

INTRODUCTION: Deep vein thrombosis (DVT) is one of the common complications of orthopedic surgery. Low molecular weight heparin (LMWH) is a usually used agent for DVT, but it would increase the risk of bleeding. LRRFIP1 has been shown to play an important role in the formation of thrombosis. Therefore, we investigated the effect of LRRFIP1 shRNA lentivirus on DVT in mice. MATERIALS AND METHODS: Lentiviral Vectors carrying LRRFIP1 shRNA were constructed and transfected into cultured mouse bone marrow cells (BMCs). Male ICR mice were irradiated with a single dose of 9.5 Gy and then were injected with different agents through the tail vein. Stasis venous thrombosis was induced by inferior vena cava (IVC) ligation. Mice were sacrificed on the 1st, 3rd and 7th day post operation and the thrombi were removed, blotted the excess blood on it with filter paper and immediately weighed. P-selectin and d-Dimer were determined by enzyme-linked immunosorbent assay (ELISA). RESULTS: LRRFIP1 shRNA significantly suppressed the expression of LRRFIP1 in the thrombi. In contrast, low molecular weight heparin (LMWH) and negative shRNA exhibited little effect on the expression of LRRFIP1. LRRFIP1 shRNA, LMWH and negative shRNA inhibited the thrombus formation in vivo significantly. The plasma P-selectin and d-Dimer levels were significantly increased after IVC ligation. LRRFIP1 shRNA significantly decreased the plasma P-selectin and d-Dimer levels. However, LMWH and negative shRNA showed little effects on the levels of plasma P-selectin and d-Dimer. CONCLUSION: LRRFIP1 shRNA might represent a promising prevention strategy for DVT.