Biology subjects

Severino, P.

Publications and source records attributed to Severino, P..

3 recordsLinked to original sources

bioRxiv2025

SiteCELL enables on-site PBMCs purification and cryopreservation for immune single cell profiling of diverse ancestries

Single cell genomics has improved our knowledge of immune function and heterogeneity. In recent years, the steady increase in the number of cells and individuals profiled as part of large multinational projects has enabled the characterization of cellular differences across human populations and ancestries. However, methods for collecting and processing peripheral blood mononuclear cells (PBMCs) for downstream single cell sequencing are difficult to implement in remote and rural settings. This has resulted in a lack of representation of underserved communities across the Global South in current initiatives. Hence, we developed SiteCELL, a method that enables purification and cryopreservation of PBMCs from whole blood at the site of collection using minimal laboratory equipment and without electricity. By comparing matched samples of purified PBMCs, we showed that SiteCELL performs as well as ficoll density gradient (FDG), both in laboratory and rural settings. This method ensures accurate recovery of cell type proportions and excels in reducing stress and minimizing variability across sampling batches. These advantages make it particularly well suited for implementation in challenging settings across countries, thereby enabling the inclusion of underrepresented ancestries in cellular atlases.

immunology↗

bioRxiv2025

The Single-Cell Landscape of Peripheral and Tumor-infiltrating Immune Cells in HPV- HNSCC

Head and neck squamous cell carcinoma (HNSCC) is the sixth most common cancer worldwide. HPV-negative HNSCC, which arises in the upper airway mucosa, is particularly aggressive, with nearly half of patients succumbing to the disease within five years and limited response to immune checkpoint inhibitors compared to other cancers. There is a need to further explore the complex immune landscape in HPV-negative HNSCC to identify potential therapeutic targets. Here, we integrated two single-cell RNA sequencing datasets from 29 samples and nearly 300,000 immune cells to investigate immune cell dynamics across tumor progression and lymph node metastasis. Notable shifts toward adaptative immune cell populations were observed in the 14 distinct HNSCC-associated peripheral blood mononuclear (PBMCs) and 21 tumor-infiltrating immune cells (TICs) considering disease stages. All PBMCs and TICs revealed unique molecular signatures correlating with lymph node involvement; however, broadly, TICs increased ligand expression among effector cytokines, growth factors, and interferon-related genes. Pathway analysis comparing PBMCs and TICs further confirmed active cell signaling among Monocyte-Macrophage, Dendritic cell, Natural Killer (NK), and T cell populations. Receptor-ligand analysis revealed significant communication patterns shifts among TICs, between CD8+ T cells and NK cells, showing heightened immunosuppressive signaling that correlated with disease progression. In locally invasive HPV-negative HNSCC samples, highly multiplexed immunofluorescence assays highlighted peri-tumoral clustering of exhausted CD8+ T and NK cells, alongside their exclusion from intra-tumoral niches. These findings emphasize cytotoxic immune cells as valuable biomarkers and therapeutic targets, shedding light on the mechanisms by which the HNSCC sustainably evades immune responses.

cancer biology↗

bioRxiv2018

Evidence of a noncoding transcript of the RIPK2 gene overexpressed in head and neck tumor

Receptor-interacting proteins are a family of serine/threonine kinases, which integrate extra and intracellular stress signals caused by different factors, including infections, inflammation and DNA damage. Receptor-interacting serine/threonine-protein kinase 2 (RIP-2) is a member of this family and an important component of the nuclear factor NF-kappa-B signaling pathway. The corresponding human gene RIPK2 generates two transcripts by alternative splicing, the full-length and a short transcript. The short transcript has a truncated 5 sequence, which results in a predicted isoform with a partial kinase domain but able to transduce signals through its caspase recruitment domain. In this study, the expression of RIPK2 was investigated in human tissue samples and, in order to determine if both transcripts are similarly regulated at the transcriptional level, cancer cell lines were submitted to temperature and acid stresses. We observed that both transcripts are expressed in all tissues analyzed, with higher expression of the short one in tumor samples, and they are differentially regulated following temperature stress. Despite transcription, no corresponding protein for the short transcript was detected in tissues and cell lines analyzed. We propose that the shorter transcript is a noncoding RNA and that its presence in the cell may play regulatory roles and affect inflammation and other biological processes related to the kinase activity of RIP-2.

molecular biology↗