Methods of Isolation, Characterization and Expansion of Human Adipose-Derived Stem Cells (ASCs): An Overview.

Considering the increasing interest in adipose-derived stem cells (ASCs) in regenerative medicine, optimization of methods aimed at isolation, characterization, expansion and evaluation of differentiation potential is critical to ensure (a) the quality of stem cells also in terms of genetic stability; (b) the reproducibility of beneficial effects; and (c) the safety of their use. Numerous studies have been conducted to understand the mechanisms that regulate ASC proliferation, growth and differentiation, however standard protocols about harvesting and processing techniques are not yet defined. It is also important to note that some steps in the procedures of harvesting and/or processing have been reported to affect recovery and/or the physiology of ASCs. Even considering the great opportunity that the ASCs provide for the identification of novel molecular targets for new or old drugs, the definition of homogeneous preparation methods that ensure adequate quality assurance and control, in accordance with current GMPs (good manufacturing practices), is required. Here, we summarize the literature reports to provide a detailed overview of the methodological issues underlying human ASCs isolation, processing, characterization, expansion, differentiation techniques, recalling at the same time their basilar principles, advantages and limits, in particular focusing on how these procedures could affect the ASC quality, functionality and plasticity.

Involvement of NOS2 Activity on Human Glioma Cell Growth, Clonogenic Potential, and Neurosphere Generation.

Aberrant nitric oxide synthase 2 (NOS2) expression has been suggested as an interesting therapeutic target that is being implicated as a component of the molecular profile of several human malignant tumors, including glioblastoma, which is the most aggressive brain tumor with limited therapeutic options and poor prognosis. The aim of the present work was to evaluate the effect of 1400W, a specific NOS2 inhibitor, on human glioma cells in terms of clonogenic potential, proliferation, migration rate, and neurosphere generation ability. NOS2 expression was determined by Western blotting. Nitric oxide (NO) production was measured through nitrite level determination. The trypan blue exclusion test and the plate colony formation assay were performed to evaluate cell proliferation and clonogenic potential. Cell proliferation and migration ability was assessed by the in vitro wound-healing assay. Neurosphere generation in a specific stemcell medium was investigated. NOS2 was confirmed to be expressed in both the glioma cell line and a human glioma primary culture, and overexpressed in relative derived neurospheres. Experiments that aimed to evaluate the influence of 1400W on U-87 MG, T98G (glioblastoma cell lines) and primary glioma cells sustained the crucial role played by NOS2 in proliferation, colony formation, migration, and neurosphere generation, thus supporting the emerging relevance of a NOS2/NO system as a prognostic factor for glioma malignancy and recurrence.

Survivin (BIRC5): Implications in cancer therapy.

Inhibitors of Apoptosis proteins (IAPs) were discovered through experiments aimed at rescuing apoptosis in insects. Classically associated with the inhibition of apoptosis, the IAP member Survivin also regulates cell cycle progression and is an essential component of the Chromosomal Passenger Complex (CPC), responsible for chromosomal segregation. Although undetectable in most adult tissues, Survivin is expressed in Adult Stem Cells (ASCs) and plays a crucial role in their maintenance. Survivin is overexpressed in most cancers, contributing to their clonal expansion. As a result, it has been proposed as a possible anticancer target for nearly two decades. In this discussion, we will explore the rationale behind Survivin as a therapeutic target, focusing on common cancer types such as carcinomas, sarcomas, and leukemias. We will delve into the modulation of Survivin by cancer pro-survival cell signaling, the association between SNPs and tumorigenesis, and its regulation by miRNAs. Finally, we will compare cell growth, clonogenic capacity, and apoptosis, along with different strategies for Survivin inhibition, including gene expression and protein activity modulation.

Effects of Intraoperative Nerve Monitoring Techniques on Voice and Swallowing Disorders after Uncomplicated Thyroidectomy: Preliminary Report of a Bi-Institutional Prospective Study.

Background: Voice and swallowing problems are frequently associated with thyroidectomy. Intermittent nerve monitoring (i-IONM) seems to provide a positive effect in reducing its prevalence. The aim of this study was to test the hypothesis that continuous intraoperative nerve monitoring (c-IONM) may reduce the prevalence of these disorders even further than i-IONM. Methods: This 3-arm prospective bi-institutional study compared 179 consecutive patients that underwent thyroidectomy: 56 without IONM, 55 with i-IONM and 67 with c-IONM. Neck dissections and laryngeal nerve palsies were excluded. Two questionnaires (VHI-10 for voice disorders and EAT-10 for swallowing disorders; both validated for Italian language use) were administered before and 1 month after surgery. Statistical significance was analyzed by the chi-squared test. Results: After thyroidectomy, no statistically significant differences were found in the three groups concerning EAT-10. although these symptoms seemed to be influenced by gastro-esophageal reflux. VHI-10 worsened in the "no-IONM" group compared with both i-IONM (p < 0.09, not quite statistically significant) and c-IONM (p < 0.04). Conclusion: Both i- and c-IONM improve voice quality independently of laryngeal nerve integrity. Reduced dissection and particularly restrained manipulation could explain these results, being particularly favorable for c-IONM.