Generation and characterization of two fibroblast-derived Baboon induced pluripotent stem cell lines.

Cross-species comparisons studying primate pluripotent stem cells and their derivatives are crucial to better understand the molecular and cellular mechanisms behind human disease and development. Within this context, Baboons (Papio anubis) have emerged as a prominent primate model for such investigations. Herein, we reprogrammed skin fibroblasts of one male individual and generated two induced pluripotent stem cell (iPSC) lines, which exhibit the characteristic ESC-like morphology, demonstrated robust expression of key pluripotency factors and displayed multilineage differentiation potential. Notably, both iPSC lines can be cultured under feeder-free conditions in commercially available medium, enhancing their value for cross-species comparisons.

Generation and characterization of two Vervet monkey induced pluripotent stem cell lines derived from fibroblasts.

Cross-species comparisons using pluripotent stem cells from primates are crucial to better understand human biology, disease, and evolution. The Vervet monkey (Chlorocebus aethiops sabaeus) serves as an important primate model for such studies, and therefore we reprogrammed skin fibroblasts derived from a male and a female individual, resulting in two induced pluripotent stem cell lines (iPSCs). These iPSCs display the characteristic ESC-like colony morphology, express key pluripotency markers, and possess the ability to differentiate into cells representing all three germ layers. Importantly, both generated cell lines can be maintained in feeder-free culture conditions using commercially available medium.

Generation and characterization of three fibroblast-derived Rhesus Macaque induced pluripotent stem cell lines.

Cross-species comparisons using pluripotent stem cells from primates are crucial to better understand human biology, disease, and evolution. An important primate model is the Rhesus macaque (Macaca mulatta), and we reprogrammed skin fibroblasts from a male individual to generate three induced pluripotent stem cell (iPSC) lines. These cells exhibit the typical ESC-like colony morphology, express common pluripotency markers, and can differentiate into cells of the three germ layers. All generated iPSC lines can be cultured under feeder-free conditions in commercially available medium and are therefore valuable resources for cross-species comparisons.

The effect of background noise and its removal on the analysis of single-cell expression data.

BACKGROUND: In droplet-based single-cell and single-nucleus RNA-seq experiments, not all reads associated with one cell barcode originate from the encapsulated cell. Such background noise is attributed to spillage from cell-free ambient RNA or barcode swapping events. RESULTS: Here, we characterize this background noise exemplified by three scRNA-seq and two snRNA-seq replicates of mouse kidneys. For each experiment, cells from two mouse subspecies are pooled, allowing to identify cross-genotype contaminating molecules and thus profile background noise. Background noise is highly variable across replicates and cells, making up on average 3-35% of the total counts (UMIs) per cell and we find that noise levels are directly proportional to the specificity and detectability of marker genes. In search of the source of background noise, we find multiple lines of evidence that the majority of background molecules originates from ambient RNA. Finally, we use our genotype-based estimates to evaluate the performance of three methods (CellBender, DecontX, SoupX) that are designed to quantify and remove background noise. We find that CellBender provides the most precise estimates of background noise levels and also yields the highest improvement for marker gene detection. By contrast, clustering and classification of cells are fairly robust towards background noise and only small improvements can be achieved by background removal that may come at the cost of distortions in fine structure. CONCLUSIONS: Our findings help to better understand the extent, sources and impact of background noise in single-cell experiments and provide guidance on how to deal with it.

A non-invasive method to generate induced pluripotent stem cells from primate urine.

Comparing the molecular and cellular properties among primates is crucial to better understand human evolution and biology. However, it is difficult or ethically impossible to collect matched tissues from many primates, especially during development. An alternative is to model different cell types and their development using induced pluripotent stem cells (iPSCs). These can be generated from many tissue sources, but non-invasive sampling would decisively broaden the spectrum of non-human primates that can be investigated. Here, we report the generation of primate iPSCs from urine samples. We first validate and optimize the procedure using human urine samples and show that suspension- Sendai Virus transduction of reprogramming factors into urinary cells efficiently generates integration-free iPSCs, which maintain their pluripotency under feeder-free culture conditions. We demonstrate that this method is also applicable to gorilla and orangutan urinary cells isolated from a non-sterile zoo floor. We characterize the urinary cells, iPSCs and derived neural progenitor cells using karyotyping, immunohistochemistry, differentiation assays and RNA-sequencing. We show that the urine-derived human iPSCs are indistinguishable from well characterized PBMC-derived human iPSCs and that the gorilla and orangutan iPSCs are well comparable to the human iPSCs. In summary, this study introduces a novel and efficient approach to non-invasively generate iPSCs from primate urine. This will extend the zoo of species available for a comparative approach to molecular and cellular phenotypes.

TAMEP are brain tumor parenchymal cells controlling neoplastic angiogenesis and progression.

Aggressive brain tumors like glioblastoma depend on support by their local environment and subsets of tumor parenchymal cells may promote specific phases of disease progression. We investigated the glioblastoma microenvironment with transgenic lineage-tracing models, intravital imaging, single-cell transcriptomics, immunofluorescence analysis as well as histopathology and characterized a previously unacknowledged population of tumor-associated cells with a myeloid-like expression profile (TAMEP) that transiently appeared during glioblastoma growth. TAMEP of mice and humans were identified with specific markers. Notably, TAMEP did not derive from microglia or peripheral monocytes but were generated by a fraction of CNS-resident, SOX2-positive progenitors. Abrogation of this progenitor cell population, by conditional Sox2-knockout, drastically reduced glioblastoma vascularization and size. Hence, TAMEP emerge as a tumor parenchymal component with a strong impact on glioblastoma progression.