RESUMEN
In this work, we have analyzed the transcriptomic changes in the brainstem of male Wistar rats 2 h after an acute stress exposure. We performed duplex-specific nuclease normalization of cDNA libraries and compared the results back-to-back for the first time. Based on our RNAseq data, we selected reference genes for RT-qPCR that are best suited for acute stress experiments. Most genes were upregulated. We detected a massive shift in neuropeptide Crh, Trh,Cga, Tshb, Uts2b, Tac4, Lep and neuropeptide receptor Hcrtr1, Sstr5, Bdkrb2, Crhr2 signaling, as well as glutamate Grin3b, Grm2 and GABA Gpr156, acetylcholine Chrm4,Chrne, adrenergic Adra2b receptors expression. A strong increase in the expression of intermediate filaments Krt83/Krt86/Krt80/Krt84/Krt87/Krt4/Krt76 and motor proteins Myo7a, Klc3 was detected. Remarkably, in the absence of astrocyte activation, we also observed signs of microglial activation at this time point. Both expression of anti-inflammatory cytokines Il13, Ccl24 and pro-inflammatory cytokine receptors Il9r, Il12rb1, Tnfrsf14, Tnfrsf13c, Tnfrsf25, Tnfrsf1b were increased. In the Wnt signaling pathway, we observed increased expression of ligands-receptors Wnt1, Wnt11, Ror2 and also negative regulators Notum, Sfrp5, Sost. RNAseq results after DSN treatment correlated at a high level with RNAseq results without DSN, but there was a proportion of genes that shifted their logFC values. They are mostly rare transcripts TPM 1-10 with higher 0.5-0.9 GC content.
Asunto(s)
Tronco Encefálico , Ratas Wistar , Transcriptoma , Animales , Masculino , Ratas , Tronco Encefálico/metabolismo , Perfilación de la Expresión Génica , Endonucleasas/metabolismo , Endonucleasas/genética , Estrés FisiológicoRESUMEN
Single-cell RNA sequencing (scRNA-seq) is a revolutionary tool for studying the physiology of normal and pathologically altered tissues. This approach provides information about molecular features (gene expression, mutations, chromatin accessibility, etc.) of cells, opens up the possibility to analyze the trajectories/phylogeny of cell differentiation and cell-cell interactions, and helps in discovery of new cell types and previously unexplored processes. From a clinical point of view, scRNA-seq facilitates deeper and more detailed analysis of molecular mechanisms of diseases and serves as a basis for the development of new preventive, diagnostic, and therapeutic strategies. The review describes different approaches to the analysis of scRNA-seq data, discusses the advantages and disadvantages of bioinformatics tools, provides recommendations and examples of their successful use, and suggests potential directions for improvement. We also emphasize the need for creating new protocols, including multiomics ones, for the preparation of DNA/RNA libraries of single cells with the purpose of more complete understanding of individual cells.
Asunto(s)
Perfilación de la Expresión Génica , ARN , Perfilación de la Expresión Génica/métodos , ARN/genética , Diferenciación Celular , Biblioteca de Genes , Análisis de Secuencia de ARN/métodosRESUMEN
Despite considerable advances in the evolution of anticancer therapies, metastasis still remains the main cause of cancer mortality. Therefore, current strategies for cancer cure should be redirected towards prevention of metastasis. Targeting metastatic pathways represents a promising therapeutic opportunity aimed at obstructing tumor cell dissemination and metastatic colonization. In this review, we focus on preclinical studies and clinical trials over the last five years that showed high efficacy in suppressing metastasis through targeting lymph node dissemination, tumor cell extravasation, reactive oxygen species, pre-metastatic niche, exosome machinery, and dormancy.
Asunto(s)
Metástasis de la Neoplasia , Neoplasias , Humanos , Neoplasias/patología , Metástasis de la Neoplasia/diagnósticoRESUMEN
Circulating tumor cells and hybrid cells formed by the fusion of tumor cells with normal cells are leading players in metastasis and have prognostic relevance. This study applies single-cell RNA sequencing to profile CD45-negative and CD45-positive circulating epithelial cells (CECs) in nonmetastatic breast cancer patients. CECs are represented by transcriptionally-distinct populations that include both aneuploid and diploid cells. CD45- CECs are predominantly aneuploid, but one population contained more diploid than aneuploid cells. CD45+ CECs mostly diploid: only two populations have aneuploid cells. Diploid CD45+ CECs annotated as different immune cells, surprisingly harbored many copy number aberrations, and positively correlated to tumor grade. It is noteworthy that cancer-associated signaling pathways areabundant only in one aneuploid CD45- CEC population, which may represent an aggressive subset of circulating tumor cells. Thus, CD45- and CD45+ CECs are highly heterogeneous in breast cancer patients and include aneuploid cells, which are most likely circulating tumor and hybrid cells, respectively, and diploid cells. DNA ploidy analysis can be an effective instrument for identifying tumor and hybrid cells among CECs. Further follow-up study is needed to determine which subsets of circulating tumor and hybrid cells contribute to breast cancer metastasis.
Asunto(s)
Neoplasias de la Mama , Células Neoplásicas Circulantes , Humanos , Femenino , Neoplasias de la Mama/genética , Neoplasias de la Mama/patología , Células Neoplásicas Circulantes/patología , Células Epiteliales/patología , Aneuploidia , Células Híbridas/patologíaRESUMEN
Metastasis is the main cause of cancer death. Metastatic foci are derived from tumor cells that detach from the primary tumor and then enter the circulation. Circulating tumor cells (CTCs) are generally associated with a high probability of distant metastasis and a negative prognosis. Most CTCs die in the bloodstream, and only a few cells form metastases. Such metastatic CTCs have a stem-like and hybrid epithelial-mesenchymal phenotype, can avoid immune surveillance, and show increased therapy resistance. Targeting metastatic CTCs and their progenitors in primary tumors and their descendants, particularly disseminated tumor cells, represents an attractive strategy for metastasis prevention. However, current therapeutic strategies mainly target the primary tumor and only indirectly affect metastasis-initiating cells. Here, we consider potential methods for preventing metastasis based on targeting molecular and cellular features of metastatic CTCs, including CTC clusters. Also, we emphasize current knowledge gaps in CTC biology that should be addressed to develop highly effective therapeutics and strategies for metastasis suppression.
Asunto(s)
Células Neoplásicas Circulantes , Biomarcadores de Tumor , Movimiento Celular , Transición Epitelial-Mesenquimal , Humanos , Metástasis de la Neoplasia , Células Neoplásicas Circulantes/patologíaRESUMEN
Despite advances in diagnostics and therapy of non-small cell lung cancer (NSCLC), the problem of prognosis and prevention of tumor progression is still highly important. Even if NSCLC is diagnosed in the early stages, almost a quarter of patients develop relapse; most of them die from recurrent disease. A large number of different markers have been proposed to predict the risk of NSCLC progression; however, none of them are used in clinical practice. It is obvious that this situation is related to the economic and methodological complexity of the proposed markers and/or their insufficient efficiency due to a lack of effective study models and tumor heterogeneity. Another reason may be that potential markers are developed for NSCLC progression in general, which is represented by at least four pathogenetically-distinct processes: synchronous lymph node metastasis, local, regional, and distant recurrence. In this review, we summarize data from published literature on clinicopathological, genetic, and molecular factors associated with different types of NSCLC progression and emphasize challenges and approaches to developing prognostic factors. In conclusion, we highlight the importance of further studies to reveal molecular mechanisms of NSCLC progression and the need for differential analysis of markers of local, regional, and distant recurrences for disease prognosis.