To assess in patients with systemic sclerosis (SSc) the concentration of different subpopulations of circulating extracellular vesicles (EVs) and their association with the progression of interstitial lung disease (PF-ILD).
The prospective study included 59 SSc cases, 54% with interstitial lung disease (ILD). Plasma levels of EVs were analysed with flow cytometry and labelled as endothelial (EEVs), platelet (PEVs), leucocyte (LEVs), and EVs, expressing ICAM1, TF, or HMGB1. The presence of ILD was defined by HRCT. Lung functional tests were done every 3-6 months over a 3-year follow-up period. PF-ILD was defined as ≥10% decline of FVC % from baseline, or ≥5-9% along with a decline in DLCO of ≥15%.
At baseline, 32/59 SSc patients had ILD, with a median disease duration of 3 years, and 38% were therapy naïve. In ILD patients, increased levels of all investigated EVs were found in respect to SSc patients without ILD (p < 0.05). Therapy naïve ILD cases had altered only ICAM1 + EVs compared to treated (p < 0.05). Multivariate regression analysis (MR) showed an independent association of PEVs (OR 1.004, 95% CI 1.001-1.01) and ICAM1 + EVs (OR 1.3, 95% CI 1.1-1.5) with ILD. During the follow-up period, 12/32 ILD patients developed PF-ILD, and in this group, the levels of all explored EVs were elevated compared to those without PF-ILD (p < 0.05). In an ROC analysis, all EVs showed a good ability to identify PF-ILD patients (p < 0.05). Cox MR confirmed the independent predictive value of ICAM1 + EVs (HR 1.1, 95% CI 1.01-1.1) with SSc PF-ILD.
Circulating EV levels are increased in SSc and correlate with ILD. In particular, ICAM1 + EVs may be a novel biomarker of PF-ILD, identifying SSc patients at high risk of progression who may require early aggressive treatment. Based on our results, the role of EVs in the pathogenesis and progression of ILD should be investigated further.
Copyright © 2025 Colic, Pruner, Damjanov, Antovic, Sefik-Bukilica, Cerinic and Antovic.

Production of specialized proresolving mediators (SPMs) during the resolution phase in the acute inflammatory response is key to orchestrating complete resolution. Here, we uncovered a trihydroxy resolvin in fresh human saliva. We identified and determined its complete stereochemistry as 7S,16R,17R-trihydroxy-4Z,8E,10Z,12E,14E,19Z-docosahexaenoic acid (17R-RvD2) using total organic synthesis and matching of physical properties. The 17R-RvD2 was produced by activated human M2-like macrophages, M1-like macrophages, and human peripheral blood monocytes. 17R-RvD2 displayed potent proresolving functions (picomolar to nanomolar). Topical application of 17R-RvD2 on mouse ear skin reduced neutrophilic infiltration (~50%). 17R-RvD2 increased M2 markers CD206 and CD163 on human monocyte-derived macrophages and enhanced efferocytosis of senescent red blood cells by M2-like macrophages (EC50 ~ 2.6 × 10-14 M). In addition, 17R-RvD2 activated the RvD2 receptor and was equipotent to its epimer RvD2. 17R-RvD2 also significantly increased phagocytosis of Escherichia coli by human neutrophils. Together, these results establish the complete stereochemistry and potent proresolving functions of the previously unknown 17R-RvD2.

Cross-linked polymer blends from natural compounds, namely gelatin (Gel), chitosan (CS), and synthetic poly (vinyl alcohol) (PVA), have received increasing scrutiny because of their versatility, biocompatibility, and ease of use for tissue engineering. Previously, Gel/CS/PVA [1:1:1] hydrogel produced via the freeze-drying process presented enhanced mechanical properties. This study aimed to investigate the biocompatibility and chondrogenic potential of a steam-sterilized Gel/CS/PVA hydrogel using differentiation of human adipose-derived mesenchymal stromal cells (AD-hMSC) and cartilage marker expression. AD-hMSC displayed fibroblast-like morphology, 90% viability, and 69% proliferative potential. Mesenchymal profiles CD73 (98.3%), CD90 (98.6%), CD105 (97.0%), CD34 (1.11%), CD45 (0.27%), HLA-DR (0.24%); as well as multilineage potential, were confirmed. Chondrogenic differentiation of AD-hMSC in monolayer revealed the formation of cartilaginous nodules composed of glycosaminoglycans after 21 days. Compared to nonstimulated cells, hMSC-derived chondrocytes shifted the expression of CD49a from 2.82% to 40.6%, CD49e from 51.4% to 92.2%, CD54 from 9.66 to 37.2%, and CD151 from 45.1% to 75.8%. When cultured onto Gel/CS/PVA hydrogel during chondrogenic stimulation, AD-hMSC changed to polygonal morphology, and chondrogenic nodules increased by day 15, six days earlier than monolayer-differentiated cells. SEM analysis showed that hMSC-derived chondrocytes adhered to the surface with extended filopodia and abundant ECM formation. Chondrogenic nodules were positive for aggrecan and type II collagen, two of the most abundant components in cartilage. This study supports the biocompatibility of AD-hMSC onto steam-sterilized GE/CS/PVA hydrogels and its improved potential for chondrocyte differentiation. Hydrogel properties were not altered after steam sterilization, which is relevant for biosafety and biomedical purposes.

Cytotoxic CD8 +T lymphocytes (CTLs) are key players of adaptive anti-tumor immunity based on their ability to specifically recognize and destroy tumor cells. Many cancer immunotherapies rely on unleashing CTL function. However, tumors can evade killing through strategies which are not yet fully elucidated. To provide deeper insight into tumor evasion mechanisms in an antigen-dependent manner, we established a human co-culture system composed of tumor and primary immune cells. Using this system, we systematically investigated intrinsic regulators of tumor resistance by conducting a complementary CRISPR screen approach. By harnessing CRISPR activation (CRISPRa) and CRISPR knockout (KO) technology in parallel, we investigated gene gain-of-function as well as loss-of-function across genes with annotated function in a colon carcinoma cell line. CRISPRa and CRISPR KO screens uncovered 187 and 704 hits, respectively, with 60 gene hits overlapping between both. These data confirmed the role of interferon-γ (IFN-γ), tumor necrosis factor α (TNF-α) and autophagy pathways and uncovered novel genes implicated in tumor resistance to killing. Notably, we discovered that ILKAP encoding the integrin-linked kinase-associated serine/threonine phosphatase 2 C, a gene previously unknown to play a role in antigen specific CTL-mediated killing, mediate tumor resistance independently from regulating antigen presentation, IFN-γ or TNF-α responsiveness. Moreover, our work describes the contrasting role of soluble and membrane-bound ICAM-1 in regulating tumor cell killing. The deficiency of membrane-bound ICAM-1 (mICAM-1) or the overexpression of soluble ICAM-1 (sICAM-1) induced resistance to CTL killing, whereas PD-L1 overexpression had no impact. These results highlight the essential role of ICAM-1 at the immunological synapse between tumor and CTL and the antagonist function of sICAM-1.
© 2023, Herzfeldt et al.

Colorectal cancer (CRC) is a leading cause of cancer-associated death. In the tumor site, the interplay between effector immune cells and cancer cells determines the balance between tumor elimination or outgrowth. We discovered that the protein TMEM123 is over-expressed in tumour-infiltrating CD4 and CD8 T lymphocytes and it contributes to their effector phenotype. The presence of infiltrating TMEM123+ CD8+ T cells is associated with better overall and metastasis-free survival. TMEM123 localizes in the protrusions of infiltrating T cells, it contributes to lymphocyte migration and cytoskeleton organization. TMEM123 silencing modulates the underlying signaling pathways dependent on the cytoskeletal regulator WASP and the Arp2/3 actin nucleation complex, which are required for synaptic force exertion. Using tumoroid-lymphocyte co-culture assays, we found that lymphocytes form clusters through TMEM123, anchoring to cancer cells and contributing to their killing. We propose an active role for TMEM123 in the anti-cancer activity of T cells within tumour microenvironment.
Copyright © 2023 Pesce, Cordiglieri, Bombaci, Eppenberger-Castori, Oliveto, Manara, Crosti, Ercan, Coto, Gobbini, Campagnoli, Donnarumma, Martinelli, Bevilacqua, De Camilli, Gruarin, Sarnicola, Cassinotti, Baldari, Viale, Biffo, Abrignani, Terracciano and Grifantini.