[Bipolar Action of Inhibitor of Vasculogenic Mimicry on Gene Expression in Melanoma Cells].

Multiple exogenous or endogenous factors alter gene expression patterns by different mechanisms that are poorly understood. We used RNA-Seq analysis in order to study changes in gene expression in melanoma cells that are capable of vasculogenic mimicry that is inhibited upon the action of an inhibitor of vasculogenic mimicry. Here, we show that the drug induces a strong upregulation of 50 genes that control the cell cycle and microtubule cytoskeleton coupled with a strong downregulation of 50 genes that control different cellular metabolic processes. We found that both groups of genes are simultaneously regulated by multiple sets of transcription factors. We conclude that one way for coordinated regulation of large groups of genes is regulation simultaneously by multiple transcription factors.

TCAF2 drives glioma cellular migratory/invasion properties through STAT3 signaling.

Glioma is an intracranial tumor characterized by high mortality and recurrence rates. In the present study, the association of TRPM8 channel-associated factor 2 (TCAF2) in glioma was investigated using bioinformatics, showing significant relationships with age, WHO grade, IDH, and 1p/19q status, as well as being an independent predictor of prognosis. Immunohistochemistry of a glioma sample microarray showed markedly increased TCAF2 expression in glioblastoma relative to lower-grade glioma, with elevated expression predominating in the tumor center. Raised TCAF2 levels promote glioma cell migratory/invasion properties through the epithelial-to-mesenchymal transition-like (EMT-like) process, shown by Transwell and scratch assays and western blotting. It was further found that the effects of TCAF2 were mediated by the activation of STAT3. These results suggest that TCAF2 promotes glioma cell migration and invasion, rendering it a potential drug target in glioma therapy.

The Association between TRP Channels Expression and Clinicopathological Characteristics of Patients with Pancreatic Adenocarcinoma.

Pancreatic adenocarcinoma (PDAC) has low survival rates worldwide due to its tendency to be detected late and its characteristic desmoplastic reaction, which slows the use of targeted therapies. As such, the discovery of new connections between genes and the clinicopathological parameters contribute to the search for new biomarkers or targets for therapy. Transient receptor potential (TRP) channels are promising tools for cancer therapy or markers for PDAC. Therefore, in this study, we selected several genes encoding TRP proteins previously reported in cellular models, namely, Transient Receptor Potential Cation Channel Subfamily V Member 6 (TRPV6), Transient receptor potential ankyrin 1 (TRPA1), and Transient receptor potential cation channel subfamily M (melastatin) member 8 (TRPM8), as well as the TRPM8 Channel Associated Factor 1 (TCAF1) and TRPM8 Channel Associated Factor 2 (TCAF2) proteins, as regulatory factors. We analyzed the expression levels of tumors from patients enrolled in public datasets and confirmed the results with a validation cohort of PDAC patients enrolled in the Clinical Institute Fundeni, Romania. We found significantly higher expression levels of TRPA1, TRPM8, and TCAF1/F2 in tumoral tissues compared to normal tissues, but lower expression levels of TRPV6, suggesting that TRP channels have either tumor-suppressive or oncogenic roles. The expression levels were correlated with the tumoral stages and are related to the genes involved in calcium homeostasis (Calbindin 1 or S100A4) or to proteins participating in metastasis (PTPN1). We conclude that the selected TRP proteins provide new insights in the search for targets and biomarkers needed for therapeutic strategies for PDAC treatment.

Partners in Crime: Towards New Ways of Targeting Calcium Channels.

The characterization of calcium channel interactome in the last decades opened a new way of perceiving ion channel function and regulation. Partner proteins of ion channels can now be considered as major components of the calcium homeostatic mechanisms, while the reinforcement or disruption of their interaction with the channel units now represents an attractive target in research and therapeutics. In this review we will focus on the targeting of calcium channel partner proteins in order to act on the channel activity, and on its consequences for cell and organism physiology. Given the recent advances in the partner proteins' identification, characterization, as well as in the resolution of their interaction domain structures, we will develop the latest findings on the interacting proteins of the following channels: voltage-dependent calcium channels, transient receptor potential and ORAI channels, and inositol 1,4,5-trisphosphate receptor.

Staged Regenerative-Phenotype Modification Therapy for Multiple RT3 Gingival Recessions in Periodontally Compromised Anterior Mandibular Teeth: A Case Report.

We report the successful treatment of multiple recession type (RT) 3 gingival recessions in periodontally compromised mandibular anterior teeth with limited keratinized tissue. A 35-yearold man with stage III, grade C periodontitis underwent a two-stage intervention. Initially, a modification of the connective tissue graft (m-CTG) wall technique was used as part of phenotype modification therapy. The CTG acted as a protective 'wall,' securing space for periodontal regeneration, enhancing root coverage, soft tissue thickness, and keratinized mucosal width. Recombinant human fibroblast growth factor-2 and carbonate apatite promoted periodontal regeneration. This procedure successfully facilitated periodontal regeneration, resulting in the transition from RT3 to RT2 gingival recession and adequate keratinized mucosal width. Eighteen months later, the second surgery used a tunneled coronally advanced flap (TCAF) for root coverage. TCAF involved combining a coronally advanced flap and tunnel technique by elevating the trapezoidal surgical papilla and using a de-epithelialized CTG inserted beneath the tunneled flap. Root conditioning with ethylenediaminetetraacetic acid and enamel matrix derivative gel application were performed. Consequently, mean CAL gain was 5.3 mm, mean root coverage was 4.5 mm in height, and the gingival phenotype improved at the treated sites by the 12-month follow-up. This staged approach addresses the challenges of treating RT3 gingival recession with promising outcomes.

When is an SNP not an SNP?

Genomic duplications are important sources of structural change and gene innovation. In humans, the most recent and highly identical sequences (>90% homology, >1 kb long) are known as segmental duplications (SDs). Single-nucleotide variants or single-nucleotide polymorphisms within SDs have not been systematically assessed due to limitations around mapping short-read sequencing data. Single-nucleotide variant rs62486260 was flagged in a study of familial renal stone disease but it was unclear whether it was real or an artifact resulting from the presence of a SD. We describe in silico and wet-lab approaches to investigate this, using segment-specific long-PCR assays, followed by short PCR for Sanger sequencing. Our conclusion was that rs62486260 is an artifact. Our approach can be generalized to deal with other such situations.

The method described includes a two-step procedure for determining whether an apparent single-nucleotide polymorphism may be an artifact resulting from the presence of a duplicated genomic region/pseudogene. Step one involves identifying sequence differences between the two duplicated regions and designing a long PCR assay to specifically amplify each region separately. Step 2 involves amplifying a short PCR product which flanks the single-nucleotide polymorphism of interest, from the long products generated in step 1.

TCAF2 is associated with the immune microenvironment, promotes pathogenesis, and impairs prognosis in glioma.

PURPOSE: Glioma is the most common primary intracranial tumor and exhibits rapid growth and aggressiveness. TRPM8 channel-associated factor 2 (TCAF2), located in cell junctions and the plasma membrane, plays a key role in the pathogeneses of several cancers in humans. However, the role of TCAF2 in glioma has been elusive. METHODS: A combination of bioinformatic analysis using The Cancer Genome Atlas database and biological experiments, including 5-ethynyl-2'-deoxyuridine, transwell, and immunohistochemistry assays and xenotransplantation, was performed to analyze the expression level of TCAF2 and to mechanistically explore the relationship of TCAF2 with malignancy, prognosis, and the immune microenvironment in glioma. RESULTS: TCAF2 was upregulated in glioma, and its expression level correlated with tumor grade and clinical outcome. The role of TCAF2 in promoting glioma malignancy was characterized through in vitro and in vivo experiments. Additionally, we observed that TCAF2 can modulate the metabolic pathways and immune microenvironment. CONCLUSION: TCAF2 acts as an oncogene and may serve as a therapeutic target and prognostic marker in glioma.

TCAF2 in Pericytes Promotes Colorectal Cancer Liver Metastasis via Inhibiting Cold-Sensing TRPM8 Channel.

Hematogenous metastasis is the main approach for colorectal cancer liver metastasis (CRCLM). However, as the gatekeepers in the tumor vessels, the role of TPCs in hematogenous metastasis remains largely unknown, which may be attributed to the lack of specific biomarkers for TPC isolation. Here, microdissection combined with a pericyte medium-based approach is developed to obtain TPCs from CRC patients. Proteomic analysis reveals that TRP channel-associated factor 2 (TCAF2), a partner protein of the transient receptor potential cation channel subfamily M member 8 (TRPM8), is overexpressed in TPCs from patients with CRCLM. TCAF2 in TPCs is correlated with liver metastasis, short overall survival, and disease-free survival in CRC patients. Gain- and loss-of-function experiments validate that TCAF2 in TPCs promotes tumor cell motility, epithelial-mesenchymal transition (EMT), and CRCLM, which is attenuated in pericyte-conditional Tcaf2-knockout mice. Mechanistically, TCAF2 inhibits the expression and activity of TRPM8, leading to Wnt5a secretion in TPCs, which facilitates EMT via the activation of the STAT3 signaling pathway in tumor cells. Menthol, a TRPM8 agonist, significantly suppresses Wnt5a secretion in TPCs and CRCLM. This study reveals the previously unidentified pro-metastatic effects of TPCs from the perspective of cold-sensory receptors, providing a promising diagnostic biomarker and therapeutic target for CRCLM.