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Articular cartilage is a skeletal tissue of avascular nature and limited self-repair capacity. Cartilage-degenerative diseases, such as osteoarthritis (OA), are difficult to treat and often necessitate joint replacement surgery. Cartilage is a tough but flexible material and relatively easy to damage. It is, therefore, of high interest to develop methods allowing chondrocytes to recolonize, to rebuild the cartilage and to restore joint functionality. Here we studied the in vitro production of cartilage-like tissue using human articular chondrocytes exposed to the Random Positioning Machine (RPM), a device to simulate certain aspects of microgravity on Earth. To screen early adoption reactions of chondrocytes exposed to the RPM, we performed quantitative real-time PCR analyses after 24 h on chondrocytes cultured in DMEM/F-12. A significant up-regulation in the gene expression of IL6, RUNX2, RUNX3, SPP1, SOX6, SOX9, and MMP13 was detected, while the levels of IL8, ACAN, PRG4, ITGB1, TGFB1, COL1A1, COL2A1, COL10A1, SOD3, SOX5, MMP1, and MMP2 mRNAs remained unchanged. The STRING (Search Tool for the Retrieval of Interacting Genes/Proteins) analysis demonstrated among others the importance of these differentially regulated genes for cartilage formation. Chondrocytes grown in DMEM/F-12 medium produced three-dimensional (3D) spheroids after five days without the addition of scaffolds. On day 28, the produced tissue constructs reached up to 2 mm in diameter. Using specific chondrocyte growth medium, similar results were achieved within 14 days. Spheroids from both types of culture media showed the typical cartilage morphology with aggrecan positivity. Intermediate filaments form clusters under RPM conditions as detected by vimentin staining after 7 d and 14 d. Larger meshes appear in the network in 28-day samples. Furthermore, they were able to form a confluent chondrocyte monolayer after being transferred back into cell culture flasks in 1 g conditions showing their suitability for transplantation into joints. Our results demonstrate that the cultivation medium has a direct influence on the velocity of tissue formation and tissue composition. The spheroids show properties that make them interesting candidates for cellular cartilage regeneration approaches in trauma and OA therapy.

Breast cancer is the leading cause of cancer death in females. The incidence has risen dramatically during recent decades. Dismissed as an "unsolved problem of the last century", breast cancer still represents a health burden with no effective solution identified so far. Microgravity (µg) research might be an unusual method to combat the disease, but cancer biologists decided to harness the power of µg as an exceptional method to increase efficacy and precision of future breast cancer therapies. Numerous studies have indicated that µg has a great impact on cancer cells; by influencing proliferation, survival, and migration, it shifts breast cancer cells toward a less aggressive phenotype. In addition, through the de novo generation of tumor spheroids, µg research provides a reliable in vitro 3D tumor model for preclinical cancer drug development and to study various processes of cancer progression. In summary, µg has become an important tool in understanding and influencing breast cancer biology.

We would like to thank Bevelacqua et al [...].

The endothelin axis, recognized for its vasoconstrictive action, plays a central role in the pathology of pulmonary arterial hypertension (PAH). Treatment with approved endothelin receptor antagonists (ERAs), such as bosentan, ambrisentan, or macitentan, slow down PAH progression and relieves symptoms. Several findings have indicated that endothelin is further involved in the pathogenesis of certain other diseases, making ERAs potentially beneficial in the treatment of various conditions. In addition to PAH, this review summarizes the use and perspectives of ERAs in cancer, renal disease, fibrotic disorders, systemic scleroderma, vasospasm, and pain management. Bosentan has proven to be effective in systemic sclerosis PAH and in decreasing the development of vasospasm-related digital ulcers. The selective ERA clazosentan has been shown to be effective in preventing cerebral vasospasm and delaying ischemic neurological deficits and new infarcts. Furthermore, in the SONAR (Study Of Diabetic Nephropathy With Atrasentan) trial, the selective ERA atrasentan reduced the risk of renal events in patients with diabetes and chronic kidney disease. These data suggest atrasentan as a new therapy in the treatment of diabetic nephropathy and possibly other renal diseases. Preclinical studies regarding heart failure, cancer, and fibrotic diseases have demonstrated promising effects, but clinical trials have not yet produced measurable results. Nevertheless, the potential benefits of ERAs may not be fully realized.

Detachment and the formation of spheroids under microgravity conditions can be observed with various types of intrinsically adherent human cells. In particular, for cancer cells this process mimics metastasis and may provide insights into cancer biology and progression that can be used to identify new drug/target combinations for future therapies. By using the synthetic glucocorticoid dexamethasone (DEX), we were able to suppress spheroid formation in a culture of follicular thyroid cancer (FTC)-133 cells that were exposed to altered gravity conditions on a random positioning machine. DEX inhibited the growth of three-dimensional cell aggregates in a dose-dependent manner. In the first approach, we analyzed the expression of several factors that are known to be involved in key processes of cancer progression such as autocrine signaling, proliferation, epithelial−mesenchymal transition, and anoikis. Wnt/β-catenin signaling and expression patterns of important genes in cancer cell growth and survival, which were further suggested to play a role in three-dimensional aggregation, such as NFKB2, VEGFA, CTGF, CAV1, BCL2(L1), or SNAI1, were clearly affected by DEX. Our data suggest the presence of a more complex regulation network of tumor spheroid formation involving additional signal pathways or individual key players that are also influenced by DEX.

Prostate cancer is one of the leading causes of cancer mortality in men worldwide. An unusual but unique environment for studying tumor cell processes is provided by microgravity, either in space or simulated by ground-based devices like a random positioning machine (RPM). In this study, prostate adenocarcinoma-derived PC-3 cells were cultivated on an RPM for time periods of 3 and 5 days. We investigated the genes associated with the cytoskeleton, focal adhesions, extracellular matrix, growth, survival, angiogenesis, and metastasis. The gene expression of signaling factors of the vascular endothelial growth factor (VEGF), mitogen-activated protein kinase (MAPK), and PI3K/AKT/mTOR (PAM) pathways was investigated using qPCR. We performed immunofluorescence to study the cytoskeleton, histological staining to examine the morphology, and a time-resolved immunofluorometric assay to analyze the cell culture supernatants. When PC-3 cells were exposed to simulated microgravity (s-µg), some cells remained growing as adherent cells (AD), while most cells detached from the cell culture flask bottom and formed multicellular spheroids (MCS). After 3-day RPM exposure, PC-3 cells revealed significant downregulation of the VEGF, SRC1, AKT, MTOR, and COL1A1 gene expression in MCS, whereas FLT1, RAF1, MEK1, ERK1, FAK1, RICTOR, ACTB, TUBB, and TLN1 mRNAs were not significantly changed. ERK2 and TLN1 were elevated in AD, and FLK1, LAMA3, COL4A5, FN1, VCL, CDH1, and NGAL mRNAs were significantly upregulated in AD and MCS after 3 days. After a 5-day culture in s-µg, the PC-3 cells showed significant downregulations of VEGF mRNA in AD and MCS, and FN1, CDH1, and LAMA3 in AD and SCR1 in MCS. In addition, we measured significant upregulations in FLT1, AKT, ERK1, ERK2, LCN2, COL1A1, TUBB, and VCL mRNAs in AD and MCS, and increases in FLK1, FN1, and COL4A5 in MCS as well as LAMB2, CDH1, RAF1, MEK1, SRC1, and MTOR mRNAs in AD. FAK1 and RICTOR were not altered by s-µg. In parallel, the secretion rate of VEGFA and NGAL proteins decreased. Cytoskeletal alterations (F-actin) were visible, as well as a deposition of collagen in the MCS. In conclusion, RPM-exposure of PC-3 cells induced changes in their morphology, cytoskeleton, and extracellular matrix protein synthesis, as well as in their focal adhesion complex and growth behavior. The significant upregulation of genes belonging to the PAM pathway indicated their involvement in the cellular changes occurring in microgravity.