Extracellular vesicle (EV) production is a universal feature of metazoan cells as well as prokaryotes (bMVs - bacterial microvesicles). They are small vesicles with phospholipid membrane carrying proteins, DNA and different classes of RNAs and are heavily involved in intercellular communication acting as vectors of information to target cells. For the last decade, the interest in EV research has exponentially increased though thorough studies of their roles in various pathologies that was not previously possible due to technical limitations. This review focuses on research evaluating the role of EV production in gastrointestinal (GI) cancer development in conjunction with GI microbiota and inflammatory diseases. We also discuss recent studies on the promising role of EVs and their content as biomarkers for early diagnosis of GI cancers. The bMVs have also been implicated in the pathogenesis of GI chronic inflammatory diseases, however, possible role of bMVs in tumorigenesis remains underestimated. We propose that EVs from eukaryotic cells as well as from different microbial, fungi, parasitic species and edible plants in GI tract act as mediators of intracellular and inter-species communication, particularly facilitating tumor cell survival and multi-drug resistance. In conclusion, we suggest that matching sequences from EV proteomes (available from public databases) with known protein sequences of microbiome gut bacteria will be useful in identification of antigen mimicry between evolutionary conservative protein sequences. Using this approach we identified Bacteroides spp. pseudokinase with activation loop and homology to PDGFRα, providing a proof-of-concept strategy. We speculate that existence of microbial pseudokinase that 'mimics' PDGFRα may be related to PDGFRα and Bacteroides spp. roles in colorectal carcinogenesis that require further investigation.

Macroporous scaffolds composed of chitosan (CHI), hydroxyapatite (HA), heparin (Hep), and polyvinyl alcohol (PVA) were prepared with a glutaraldehyde (GA) cross-linker by cryogelation. Addition of PVA to the reaction mixture slowed down the formation of a polyelectrolyte complex (PEC) between CHI and Hep, which allowed more thorough mixing, and resulted in the development of the homogeneous matrix structure. Freezing of the CHI-HA-GA and PVA-Hep-GA mixture led to the formation of a non-stoichiometric PEC between oppositely charged groups of CHI and Hep, which caused further efficient immobilization of bone morphogenic protein 2 (BMP-2) possible due to electrostatic interactions. It was shown that the obtained cryogel matrix released BMP-2 and supported the differentiation of rat bone marrow mesenchymal stem cells (rat BMSCs) into the osteogenic lineage. Rat BMSCs attached to cryogel loaded with BMP-2 and expressed osteocalcin in vitro. Obtained composite cryogel with PEC may have high potential for bone regeneration and tissue engineering applications.

Wound healing assay performed with automated microscopy is widely used in drug testing, cancer cell analysis, and similar approaches. It is easy to perform, and the results are reproducible. However, it is usually used as a semi-quantitative approach because of inefficient image segmentation in transmitted light microscopy. Recently, several algorithms for wound healing quantification were suggested, but none of them was tested on a large dataset. In the current study, we develop a pipeline allowing to achieve correct segmentation of the wound edges in >95% of pictures and extended statistical data processing to eliminate errors of cell culture artifacts. Using this tool, we collected data on wound healing dynamics of 10 cell lines with 10 min time resolution. We determine that the overall kinetics of wound healing is non-linear; however, all cell lines demonstrate linear wound closure dynamics in a 6-h window between the fifth and 12th hours after scratching. We next analyzed microtubule-inhibiting drugs’, nocodazole, vinorelbine, and Taxol, action on the kinetics of wound healing in the drug concentration-dependent way. Within this time window, the measurements of velocity of the cell edge allow the detection of statistically significant data when changes did not exceed 10–15%. All cell lines show decrease in the wound healing velocity at millimolar concentrations of microtubule inhibitors. However, dose-dependent response was cell line specific and drug specific. Cell motility was completely inhibited (edge velocity decreased 100%), while in others, it decreased only slightly (not more than 50%). Nanomolar doses (10–100 nM) of microtubule inhibitors in some cases even elevated cell motility. We speculate that anti-microtubule drugs might have specific effects on cell motility not related to the inhibition of the dynamic instability of microtubules.

YAP is a downstream nuclear transcription factor of Hippo pathway which plays an essential role in development, cell growth, organ size and homeostasis. It was previously identified that elevation of YAP in genomics of genetic engineered mouse (GEM) model of prostate cancer is associated with Pten/Trp53 inactivation and ARF elevation hypothesizing the essential crosstalk of AKT/mTOR/YAP with ARF in prostate cancer. However, the detailed function and trafficking of YAP in cancer cells remains unclear. Using GEM microarray model, we found ARF dysregulates Hippo and Wnt pathways. In particular, ARF knockdown reduced non-nuclear localization of YAP which led to an increase in F-actin. Mechanistically, ARF knockdown suppressed protein turnover of β-catenin/YAP, and therefore enhanced the activity of AKT and phosphorylation of YAP. Moreover, we found tea-derived carbon dots can interact with ARF in nucleus that may further lead to the non-nuclear localization of YAP. Thus, we reported a novel crosstalk of ARF/β-catenin dysregulated YAP in Hippo pathway and a new approach to stimulate ARF-mediated signaling to inhibit nuclear YAP using nanomaterials implicating an innovative avenue for treatment of cancer.

Microtubule-targeting (MT) drugs taxanes and vinca alkaloids are widely used as chemotherapeutic agents against different tumors for more than 30 years because of their ability to block mitotic progression by disrupting the mitotic spindle and activating the spindle assembly checkpoint (SAC) for a prolonged period of time. However, responses to mitotic arrest are different-some cells die during mitotic arrest, whereas others undergo mitotic slippage and survive becoming able for proliferation. Using normal fibroblasts and several cancer cell types we determined two critical doses, T1 and T2, of mitotic inhibitors (nocodazole, Taxol, and vinorelbine). T1 is the maximal dose cells can tolerate undergoing normal division, and T2 is the minimal mitostatic dose, wherein > 90% of mitotic cells are arrested in mitosis. In all studied cell lines after treatment with mitotic inhibitors in a dose above T2 cells had entered mitosis either die or undergo mitotic slippage. We show that for all three drugs used cell death during mitotic arrest and after slippage proceeded via mitochondriadependent apoptosis. We determined two types of cancer cells: sensitive to mitotic arrest, that is, undergoing death in mitosis (DiM) frequently, and resistant to mitotic arrest, that is, undergoing mitotic slippage followed by prolonged survival. We then determined that inhibition of Bcl-xL, but not other antiapoptotic proteins of the Bcl-2 group that regulate MOMP, make resistant cells susceptible to DiM induced by mitotic inhibitors. Combined treatment with MT drugs and highly specific Bcl-xL inhibitors A-1155643 or A-1331852 allows achieving 100% DiM in a time significantly shorter than maximal duration of mitotic arrest in all types of cultured cells tested. We further examined efficacy of sequential treatment of cultured cells using mitotic inhibitors followed by inhibitors of Bcl-xL anti-apoptotic protein and for the first time show that sensitivity to Bcl-xL inhibitors rapidly declines after mitotic slippage. Thus sequential use of mitotic inhibitors and inhibitors of Bcl-xL anti-apoptotic protein will be efficient only if the Bcl-xL inhibitor will be added before mitotic slippage occurs or soon afterward. The combined treatment proposed might be an efficient approach to anti-cancer therapy.