From in vitro to ex vivo: subcellular localization and uptake of graphene quantum dots into solid tumors.

Among various nanoparticles tested for pharmacological applications over the recent years, graphene quantum dots (GQDs) seem to be promising candidates for the construction of drug delivery systems due to their superior biophysical and biochemical properties. The subcellular fate of incorporated nanomaterial is decisive for transporting pharmaceuticals into target cells. Therefore a detailed characterization of the uptake of GQDs into different breast cancer models was performed. The demonstrated accumulation inside the endolysosomal system might be the reason for the particles' low toxicity, but has to be overcome for cytosolic or nuclear drug delivery. Furthermore, the penetration of GQDs into precision-cut mammary tumor slices was studied. These constitute a far closer to reality model system than monoclonal cell lines. The constant uptake into the depth of the tissue slices underlines the systems' potential for drug delivery into solid tumors.

Characterization of CD34+ Cells from Patients with Acute Myeloid Leukemia (AML) and Myelodysplastic Syndromes (MDS) Using a t-Distributed Stochastic Neighbor Embedding (t-SNE) Protocol.

Using multi-color flow cytometry analysis, we studied the immunophenotypical differences between leukemic cells from patients with AML/MDS and hematopoietic stem and progenitor cells (HSPCs) from patients in complete remission (CR) following their successful treatment. The panel of markers included CD34, CD38, CD45RA, CD123 as representatives for a hierarchical hematopoietic stem and progenitor cell (HSPC) classification as well as programmed death ligand 1 (PD-L1). Rather than restricting the evaluation on a 2- or 3-dimensional analysis, we applied a t-distributed stochastic neighbor embedding (t-SNE) approach to obtain deeper insight and segregation between leukemic cells and normal HPSCs. For that purpose, we created a t-SNE map, which resulted in the visualization of 27 cell clusters based on their similarity concerning the composition and intensity of antigen expression. Two of these clusters were "leukemia-related" containing a great proportion of CD34+/CD38- hematopoietic stem cells (HSCs) or CD34+ cells with a strong co-expression of CD45RA/CD123, respectively. CD34+ cells within the latter cluster were also highly positive for PD-L1 reflecting their immunosuppressive capacity. Beyond this proof of principle study, the inclusion of additional markers will be helpful to refine the differentiation between normal HSPCs and leukemic cells, particularly in the context of minimal disease detection and antigen-targeted therapeutic interventions. Furthermore, we suggest a protocol for the assignment of new cell ensembles in quantitative terms, via a numerical value, the Pearson coefficient, based on a similarity comparison of the t-SNE pattern with a reference.

Identification of nanoparticles as vesicular cargo via Airy scanning fluorescence microscopy and spatial statistics.

Many biomedical applications of nanoparticles on the cellular level require a characterisation of their subcellular distribution. Depending on the nanoparticle and its preferred intracellular compartment, this may be a nontrivial task, and consequently, the available methodologies are constantly increasing. Here, we show that super-resolution microscopy in combination with spatial statistics (SMSS), comprising the pair correlation and the nearest neighbour function, is a powerful tool to identify spatial correlations between nanoparticles and moving vesicles. Furthermore, various types of motion like for example diffusive, active or Lévy flight transport can be distinguished within this concept via suitable statistical functions, which also contain information about the factors limiting the motion, as well as regarding characteristic length scales. The SMSS concept fills a methodological gap related to mobile intracellular nanoparticle hosts and its extension to further scenarios is straightforward. It is exemplified on MCF-7 cells after exposure to carbon nanodots, demonstrating that these particles are stored predominantly in the lysosomes.

Uptake of carbon nanodots into human AML cells in comparison to primary hematopoietic cells.

Carbon nanodots (CNDs) comprise a class of next generation nanomaterials with a wide variety of potential applications. Here, we report on their uptake into primary hematopoietic cells from three normal donors and malignant cells from five patients with de novo acute myeloid leukemia (AML). A significant CND uptake was observed in all cell types of the normal and leukemic cells. Still, the uptake was significantly smaller for the CD34+ and CD33+ myeloid subsets of the malignant cell population as compared to the normal blood-derived CD34+ and CD33+ cells. For the T and B lymphoid cell populations as defined by CD3 and CD19 within the leukemic and normal samples a similar uptake was observed. The CNDs accumulate preferentially in small clusters in the periphery of the nucleus as already shown in previous studies for CD34+ progenitor/stem cells and human breast cancer cells. This particular subcellular localization could be useful for targeting the lysosomal compartment, which plays a pivotal role in the context of autophagy associated survival of AML cells. Our results demonstrate the usability of CNDs beyond their application for in vitro and in vivo fluorescence labeling or drug delivery into normal and malignant cells.

The Low Toxicity of Graphene Quantum Dots is Reflected by Marginal Gene Expression Changes of Primary Human Hematopoietic Stem Cells.

Graphene quantum dots (GQDs) are a promising next generation nanomaterial with manifold biomedical applications. For real world applications, comprehensive studies on their influence on the functionality of primary human cells are mandatory. Here, we report the effects of GQDs on the transcriptome of CD34+ hematopoietic stem cells after an incubation time of 36 hours. Of the 20 800 recorded gene expressions, only one, namely the selenoprotein W, 1, is changed by the GQDs in direct comparison to CD34+ hematopoietic stem cells cultivated without GQDs. Only a meta analysis reveals that the expression of 1171 genes is weakly affected, taking into account the more prominent changes just by the cell culture. Eight corresponding, weakly affected signaling pathways are identified, which include, but are not limited to, the triggering of apoptosis. These results suggest that GQDs with sizes in the range of a few nanometers hardly influence the CD34+ cells on the transcriptome level after 36 h of incubation, thereby demonstrating their high usability for in vivo studies, such as fluorescence labeling or delivery protocols, without strong effects on the functional status of the cells.

Relationship between chain length, disorder, and resistivity in polypyrrole films.

The effects of the polymerization temperature and of voltammetric cycling on the chain length and the resistivity of polypyrrole films are investigated. The studies provide further proof for the existence of at least two different types of polypyrrole, the so-called PPy I and PPy II. As the electropolymerization of conjugated systems in contrast to normal polymerization reactions is a fully activated process, the generation of these different types of PPy depends on experimental parameters such as temperature or formation potentials. UV-vis measurements demonstrate that PPy II comprises significantly shorter chains than PPy I (8-12 vs 32-64 units); moreover, film conductivity is found to increase with the fraction of PPy II. This fraction is changed via the polymerization temperature as well as by cyclic voltammetry, both of which can induce a metal-insulator transition. The counter-intuitive relationship between resistivity and chain length is interpreted in terms of disorder-dominated transport, in which the shorter chains of PPy II support the formation of delocalized electronic states, thereby increasing the localization length. Thus, our results are in agreement with recent broadband reflectivity measurements.

Impurity and edge roughness scattering in graphene nanoribbons: the Boltzmann approach.

The conductivity of graphene nanoribbons in the presence of bulk impurities and edge roughness is studied theoretically using the Boltzmann transport equation for quasi-one-dimensional systems. As the number of occupied subbands increases, the conductivity due to bulk impurities converges towards the two-dimensional case. It is shown that the dependence of the conductivity generated by edge roughness scattering depends in a distinctly different way on the sample parameters than the conductivity due to bulk scattering. The Boltzmann model furthermore predicts the amplitude of the edge-roughness-induced magnetoconductance dip as a function of the amplitude and the correlation length of the edge roughness.