Bruno M. Costa

Mesenchymal stem cells secretome as a modulator of the neurogenic niche: basic insights and therapeutic opportunities

Neural stem cells (NSCs) and mesenchymal stem cells (MSCs) share few characteristics apart from self-renewal and multipotency. In fact, the neurogenic and osteogenic stem cell niches derive from two distinct embryonary structures; while the later originates from the mesoderm, as all the connective tissues do, the first derives from the ectoderm. Therefore, it is highly unlikely that stem cells isolated from one niche could form terminally differentiated cells from the other. Additionally, these two niches are associated to tissues/systems (e.g., bone and central nervous system) that have markedly different needs and display diverse functions within the human body. Nevertheless they do share common features. For instance, the differentiation of both NSCs and MSCs is intimately associated with the bone morphogenetic protein family. Moreover, both NSCs and MSCs secrete a panel of common growth factors, such as nerve growth factor (NGF), glial derived neurotrophic factor (GDNF), and brain derived neurotrophic factor (BDNF), among others. But it is not the features they share but the interaction between them that seem most important, and worth exploring; namely, it has already been shown that there are mutually beneficially effects when these cell types are co-cultured in vitro. In fact the use of MSCs, and their secretome, become a strong candidate to be used as a therapeutic tool for CNS applications, namely by triggering the endogenous proliferation and differentiation of neural progenitors, among other mechanisms. Quite interestingly it was recently revealed that MSCs could be found in the human brain, in the vicinity of capillaries. In the present review we highlight how MSCs and NSCs in the neurogenic niches interact. Furthermore, we propose directions on this field and explore the future therapeutic possibilities that may arise from the combination/interaction of MSCs and NSCs.

Understanding TERT Promoter Mutations: A Common Path to Immortality.

Telomerase (TERT) activation is a fundamental step in tumorigenesis. By maintaining telomere length, telomerase relieves a main barrier on cellular lifespan, enabling limitless proliferation driven by oncogenes. The recently discovered, highly recurrent mutations in the promoter of TERT are found in over 50 cancer types, and are the most common mutation in many cancers. Transcriptional activation of TERT, via promoter mutation or other mechanisms, is the rate-limiting step in production of active telomerase. Although TERT is expressed in stem cells, it is naturally silenced upon differentiation. Thus, the presence of TERT promoter mutations may shed light on whether a particular tumor arose from a stem cell or more differentiated cell type. It is becoming clear that TERT mutations occur early during cellular transformation, and activate the TERT promoter by recruiting transcription factors that do not normally regulate TERT gene expression. This review highlights the fundamental and widespread role of TERT promoter mutations in tumorigenesis, including recent progress on their mechanism of transcriptional activation. These somatic promoter mutations, along with germline variation in the TERT locus also appear to have significant value as biomarkers of patient outcome. Understanding the precise molecular mechanism of TERT activation by promoter mutation and germline variation may inspire novel cancer cell-specific targeted therapies for a large number of cancer patients. Mol Cancer Res; 14(4); 315–23. ©2016 AACR.

Expression of histone methyltransferases as novel biomarkers for renal cell tumor diagnosis and prognostication.

Renal cell tumors (RCTs) are the most lethal of the common urological cancers. The widespread use of imaging entailed an increased detection of small renal masses, emphasizing the need for accurate distinction between benign and malignant RCTs, which is critical for adequate therapeutic management. Histone methylation has been implicated in renal tumorigenesis, but its potential clinical value as RCT biomarker remains mostly unexplored. Hence, the main goal of this study was to identify differentially expressed histone methyltransferases (HMTs) and histone demethylases (HDMs) that might prove useful for RCT diagnosis and prognostication, emphasizing the discrimination between oncocytoma (a benign tumor) and renal cell carcinoma (RCC), especially the chromophobe subtype (chRCC). We found that the expression levels of 3 genes—SMYD2, SETD3, and NO66—was significantly altered in a set of RCTs, which was further validated in a large independent cohort. Higher expression levels were found in RCTs compared to normal renal tissues (RNTs) and in chRCCs comparatively to oncocytomas. SMYD2 and SETD3 mRNA levels correlated with protein expression assessed by immunohistochemistry. SMYD2 transcript levels discriminated RCTs from RNT, with 82.1% sensitivity and 100% specificity [area under curve (AUC) = 0.959], and distinguished chRCCs from oncocytomas, with 71.0% sensitivity and 73.3% specificity (AUC = 0.784). Low expression levels of SMYD2, SETD3, and NO66 were significantly associated with shorter disease-specific and disease-free survival, especially in patients with non-organ confined tumors. We conclude that expression of selected HMTs and HDMs might constitute novel biomarkers to assist in RCT diagnosis and assessment of tumor aggressiveness.

Variant Rett syndrome in a girl with a pericentric X-chromosome inversion leading to epigenetic changes and overexpression of the MECP2 gene

Rett syndrome is a neurodevelopmental disorder caused by mutations in the MECP2 gene. We investigated the genetic basis of disease in a female patient with a Rett‐like clinical. Karyotype analysis revealed a pericentric inversion in the X chromosome −46,X,inv(X)(p22.1q28), with breakpoints in the cytobands where the MECP2 and CDKL5 genes are located. FISH analysis revealed that the MECP2 gene is not dislocated by the inversion. However, and in spite of a balanced pattern of X inactivation, this patient displayed hypomethylation and an overexpression of the MECP2 gene at the mRNA level in the lymphocytes (mean fold change: 2.55 ± 0.38) in comparison to a group of control individuals; the expression of the CDKL5 gene was similar to that of controls (mean fold change: 0.98 ± 0.10). No gains or losses were detected in the breakpoint regions encompassing known or suspected transcription regulatory elements. We propose that the de‐regulation of MECP2 expression in this patient may be due to alterations in long‐range genomic interactions caused by the inversion and hypothesize that this type of epigenetic de‐regulation of the MECP2 may be present in other RTT‐like patients.

A semiautomated microfluidic platform for real-time investigation of nanoparticles’ cellular uptake and cancer cells’ tracking

AIM Develop a platform composed of labeled dendrimer nanoparticles (NPs) and a microfluidic device for real-time monitoring of cancer cells fate. MATERIALS & METHODS Carboxymethylchitosan/poly(amidoamine) dendrimer NPs were labeled with fluorescein-5(6)-isothiocyanate and characterized using different physicochemical techniques. After, HeLa, HCT-116 and U87MG were cultured in the presence of NPs, and cell viability and internalization efficiency in static (standard culture) and dynamic (microfluidic culture) conditions were investigated. RESULTS Cancer cells cultured with NPs in dynamic conditions were viable and presented higher internalization levels as compared with static 2D cultures. CONCLUSION This work demonstrated that the proposed microfluidic-based platform allows real-time monitoring, which upon more studies, namely, the assessment of an anticancer drug release effect could be used for cancer theranostics.

SETDB2 and RIOX2 are differentially expressed among renal cell tumor subtypes, associating with prognosis and metastization

ABSTRACT Increasing detection of small renal masses by imaging techniques entails the need for accurate discrimination between benign and malignant renal cell tumors (RCTs) as well as among malignant RCTs, owing to differential risk of progression through metastization. Although histone methylation has been implicated in renal tumorigenesis, its potential as biomarker for renal cell carcinoma (RCC) progression remains largely unexplored. Thus, we aimed to characterize the differential expression of histone methyltransferases (HMTs) and histone demethylases (HDMs) in RCTs to assess their potential as metastasis biomarkers. We found that SETDB2 and RIOX2 (encoding for an HMT and an HDM, respectively) expression levels was significantly altered in RCTs; these genes were further selected for validation by quantitative RT-PCR in 160 RCTs. Moreover, SETDB2, RIOX2, and three genes encoding for enzymes involved in histone methylation (NO66, SETD3, and SMYD2), previously reported by our group, were quantified (RT-PCR) in an independent series of 62 clear cell renal cell carcinoma (ccRCC) to assess its potential role in ccRCC metastasis development. Additional validation was performed using TCGA dataset. SETDB2 and RIOX2 transcripts were overexpressed in RCTs compared to renal normal tissues (RNTs) and in oncocytomas vs. RCCs, with ccRCC and papillary renal cell carcinoma (pRCC) displaying the lowest levels. Low SETDB2 expression levels and higher stage independently predicted shorter disease-free survival. In our 62 ccRCC cohort, significantly higher RIOX2, but not SETDB2, expression levels were depicted in cases that developed metastasis during follow-up. These findings were not apparent in TCGA dataset. We concluded that SETDB2 and RIOX2 might be involved in renal tumorigenesis and RCC progression, especially in metastatic spread. Moreover, SETDB2 expression levels might independently discriminate among RCC subgroups with distinct outcome, whereas higher RIOX2 transcript levels might identify ccRCC cases with more propensity to endure metastatic dissemination.