Bone marrow derived cells in the tumour microenvironment contain cells with primitive haematopoietic phenotype

Infiltration of bone marrow derived cells is part of the angiogenic switch required for uncontrolled tumour growth. However, the nature of the tumour-infiltrating cells from bone marrow has not been fully elucidated. To investigate the phenotype of bone marrow derived cells within a tumour, we employed the Lewis lung carcinoma (LLC) murine tumour model. We followed bone marrow derivation of tumour-infiltrating cells through transplantation of CD45.2 bone marrow cells into pre-irradiated CD45.1 mice. We found robust CD45.2 donor type chimerism in bone marrow and blood of CD45.1 recipient tumour-bearing mice. Flow cytometric analysis of LLC tumours showed, in addition to previously described pro-angiogenic CD45⁺VEGFR2⁺‘endothelial progenitor cells’ (EPC), or CD45⁺Tie2⁺‘Tie2-expressing monocytes’ (TEM), incorporation of donor type lineage marker negative (Lin⁻) and Lin⁻Sca1⁺ undifferentiated haematopoietic cell types. Immunohistochemical analysis confirmed the extravasal location of the primitive haematopoietic cells. Flow-cytometric sorting of bone marrow cells and subsequent analysis in haematopoietic colony-forming assays revealed that cells with a Lin⁻Sca1⁺ phenotype, which were initially negative for VEGFR2 and Tie2, gave rise to VEGFR2⁺ and/or Tie2⁺ cells. Moreover, Lin⁻ bone marrow cells pre-labelled with the membrane dye PKH26 (a red fluorochrome) and transplanted i.v. into tumour-bearing mice were found to extravasate and incorporate into LLC tumours within 24 hrs. Thus, primitive haematopoietic precursors which are thought to be precursors of EPC and TEMs, constitute a part of the tumour microenvironment. This makes them an attractive target cell population for tumour-directed cellular therapies.     

Probing the Conformation of a Prion Protein Fibril with Hydrogen Exchange

A fragment of the prion protein, PrP(89–143, P101L), bearing a mutation implicated in familial prion disease, forms fibrils that have been shown to induce prion disease when injected intracerebrally into transgenic mice expressing full-length PrP containing the P101L mutation. In this study, we utilize amide hydrogen exchange measurements to probe the organization of the peptide in its fibrillar form. We determined the extent of hydrogen exchange first by tandem proteolysis, liquid chromatography, and mass spectrometry (HXMS) and then by exchange-quenched NMR. Although single amide resolution is afforded by NMR measurements, HXMS is well suited to the study of natural prions because it does not require labeling with NMR active isotopes. Thus, natural prions obtained from infected animals can be compared with model systems such as PrP(89–143, P101L) studied here. In our study, we find two segments of sequence that display a high level of protection from exchange, residues 102–109 and 117–136. In addition, there is a region that displays exchange behavior consistent with the presence of a conformationally heterogeneous turn. We discuss our data with respect to several structural models proposed for infectious PrP aggregates and highlight HXMS as one of the few techniques well suited to studying natural prions.     

Arachidonic acid accumulates in the stromal macrophages during thymus involution in diabetes

Diabetes is a debilitating disease with chronic evolution that affects many tissues and organs over its course. Thymus is an organ that is affected early after the onset of diabetes, gradually involuting until it loses most of its thymocyte populations. We show evidence of accumulating free fatty acids with generation of eicosanoids in the diabetic thymus and we present a possible mechanism for the involution of the organ during the disease. Young rats were injected with streptozotocin and their thymuses examined for cell death by flow cytometry and TUNEL reaction. Accumulation of lipids in the diabetic thymus was investigated by histology and electron microscopy. The identity and quantitation of accumulating lipids was done with gas chromatography-mass spectrometry and liquid chromatography. The expression and dynamics of the enzymes were monitored via immunohistochemistry. Diabetes causes thymus involution by elevating the thymocyte apoptosis. Exposure of thymocytes to elevated concentration of glucose causes apoptosis. After the onset of diabetes, there is a gradual accumulation of free fatty acids in the stromal macrophages including arachidonic acid, the substrate for eicosanoids. The eicosanoids do not cause thymocyte apoptosis but administration of a cyclooxygenase inhibitor reduces the staining for ED1, a macrophage marker whose intensity correlates with phagocytic activity. Diabetes causes thymus involution that is accompanied by accumulation of free fatty acids in the thymic macrophages. Excess glucose is able to induce thymocyte apoptosis but eicosanoids are involved in the chemoattraction of macrophage to remove the dead thymocytes.     

A novel electrospinning target to improve the yield of uniaxially aligned fibers

Electrospinning is a useful technique that can generate micro and nanometer‐sized fibers. Modification of the electrospinning parameters, such as deposition target geometry, can generate uniaxially aligned fibers for use in diverse applications ranging from tissue engineering to material fabrication. For example, meshes of fibers have been shown to mimic the extracellular matrix networks for use in smooth muscle cell proliferation. Further, aligned fibers can guide neurites to grow along the direction of the fibers. Here we present a novel electrospinning deposition target that combines the benefits of two previously reported electrodes: the standard parallel electrodes and the spinning wheel with a sharpened edge. This new target design significantly improves aligned fiber yield. Specifically, the target consists of two parallel aluminum plates with sharpened edges containing a bifurcating angle of 26°. Electric field computations show a larger probable area of aligned electric field vectors. This new deposition target allows fibers to deposit on a larger cross‐sectional area relative to the existing parallel electrode and at least doubles the yield of uniaxially aligned fibers. Further, fiber alignment and morphology are preserved after collection from the deposition target. © 2009 American Institute of Chemical Engineers Biotechnol. Prog., 2009     

Tumour‐associated fibroblasts and mesenchymal stem cells: more similarities than differences

Tumour‐associated fibroblasts (TAFs) are part of the tumour stroma, providing functional and structural support for tumour progression and development. The origin and biology of TAFs are poorly understood, but within the tumour environment, TAFs become activated and secrete different paracrine and autocrine factors involved in tumorigenesis. It has been shown that bone marrow mesenchymal stem cells (MSCs) can be recruited into the tumours, where they proliferate and acquire a TAF‐like phenotype. We attempted to determine to what extent TAFs characteristics in vitro juxtapose to MSCs’ definition, and we showed that TAFs and MSCs share immunophenotypic similarities, including the presence of certain cell surface molecules [human leukocyte antigen‐DR subregion (HLA‐DR), CD29, CD44, CD73, CD90, CD106 and CD117]; the expression of cytoskeleton and extracellular matrix proteins, such as vimentin, α‐smooth muscle actin, nestin and trilineage differentiation potential (to adipocytes, chondrocytes and osteoblasts). When compared to MSCs, production of cytokines, chemokines and growth factors showed a significant increase in TAFs for vascular endothelial growth factor, transforming growth factor‐β1, interleukins (IL‐4, IL‐10) and tumour necrosis factor α. Proliferation rate was highly increased in TAFs and fibroblast cell lines used in our study, compared to MSCs, whereas ultrastructural details differentiated the two cell types by the presence of cytoplasmic elongations, lamellar content lysosomes and intermediate filaments. Our results provide supportive evidence to the fact that TAFs derive from MSCs and could be a subset of ‘specialized’ MSCs.     

Decreasing the Effective Thermal Conductivity in Glass Supported Thermoelectric Layers

As thermoelectric devices begin to make their way into commercial applications, the emphasis is put on decreasing the thermal conductivity. In this purely theoretical study, finite element analysis is used to determine the effect of a supporting material on the thermal conductivity of a thermoelectric module. The simulations illustrate the heat transfer along a sample, consisting from Cu, Cu₂O and PbTe thermoelectric layers on a 1 mm thick Pyrex glass substrate. The influence of two different types of heating, at a constant temperature and at a constant heat flux, is also investigated. It is revealed that the presence of a supporting material plays an important role on lowering the effective thermal conductivity of the layer-substrate ensemble. By using thinner thermoelectric layers the effective thermal conductivity is further reduced, almost down to the value of the glass substrate. As a result, the temperature gradient becomes steeper for a fixed heating temperature, which allows the production of devices with improved performance under certain conditions. Based on the simulation results, we also propose a model for a robust thin film thermoelectric device. With this suggestion, we invite the thermoelectric community to prove the applicability of the presented concept for practical purposes.     

Amino acid precursors for the detection of transketolase activity in Escherichia coli auxotrophs

Probes were developed for the in vivo detection of transketolase activity by the use of a complementation assay in Escherichia coli auxotrophs They combine the d-threo ketose moiety recognised by transketolase and the side chain of leucine or methionine. These compounds were donor substrates of yeast transketolase leading to the release of the corresponding α-hydroxyaldehydes which could be converted in E. coli by a cascade of reactions into leucine or methionine required for cellular growth.     

No Conventional Function for the Conventional Kinesin?

A paper by DeGiorgis et al. (DeGiorgis JA, Petukhova TA, Evans TA, Reese TS. Kinesin-3 is an organelle motor in the squid giant axon. Traffic 2008; DOI: 10.1111/j.1600-0854.2008.00809.x) in this issue of Traffic reports on the identification and function of a second squid kinesin, a kinesin-3 motor. As expected, the newly discovered motor associates with axoplasmic organelles in situ and powers motility along microtubules of vesicles isolated from squid axoplasm. Less expected was the finding that kinesin-3 may be the predominant motor for anterograde organelle movement in the squid axon, which challenges the so far undisputed view that this function is fulfilled by the conventional kinesin, kinesin-1. These novel findings let us wonder what the real function of kinesin-1--the most abundant motor in squid axons--actually is.     

Predation risk cues associated with killed conspecifics affect the behavior and reproduction of prey animals

Organisms often perceive risk to be attacked by natural enemies through cues that accompany or persist after attacks on conspecifics. Under the risk of attack, preys may prioritize anti-enemy behaviours, sacrificing their feeding activities. These enemy-induced changes can affect the prey population growth. The presence of killed congeners is a past attack events cue but, in terrestrial prey–enemy systems, its effect on prey population biology is an open question. This was addressed here by studying both the space use and reproductive effort of aphids exposed to congeners killed by parasitoids. To test whether preys adjust their responses with the threat intensity, aphids' responses in presence of killed conspecifics were compared to those measured when directly exposed to non-consumptive parasitoids. The results showed that killed congeners are perceived by aphids and led to both changes in their space use and a strong decline in their population size. This experiment is the first to demonstrate that the presence of congeners killed by an enemy can contribute to the suppressive effects of natural enemies on prey populations.