This study explores a surface modification strategy that mimics the native cellular environment through the layer-by-layer assembly of natural polyelectrolytes. Specifically, we developed a multilayer matrix composed of 10 alternating layers of chitosan (a polycation) and hyaluronic acid (a polyanion), seeded with Wharton’s jelly mesenchymal stem cells (WJ-MSCs) derived from human umbilical cords. These cells are attractive for cartilage regeneration due to their accessibility, robust differentiation potential, and low immunogenicity. WJ-MSCs were cultured on the multilayer films at a density of 3000 cells/cm2 in standard growth medium. Positive controls included cells on multilayer films supplemented with transforming growth factor-beta (TGF-β), while negative controls were cells cultured on glass in standard medium. Cell morphology, proliferation, matrix formation, and expression of key chondrogenic markers were assessed. The WJ-MSCs adhered well, exhibited fibroblast-like morphology, and expressed characteristic MSC markers (CD44, CD90, CD73) while lacking hematopoietic markers (CD34, CD45), as defined by ISCT guidelines. The chitosan–hyaluronic acid (CHI-HA) films supported spontaneous chondrogenic differentiation, as demonstrated by upregulation of chondrogenic genes and proteins, and positive staining for chondroitin sulfate. Notably, chondrogenic differentiation on CHI-HA films enhanced the immunomodulatory profile of WJ-MSCs, as shown by upregulation of IL-10 and selective modulation of TLR expression. Despite increased TNF-α, this was attributed to TGF-β signaling rather than inflammation. Overall, CHI-HA films promoted both chondrogenic and immunoregulatory functions, offering a promising platform for cartilage tissue engineering.

INTRODUCTION:Electronic cigarettes (e-cigarettes) are promoted as safer alternatives to combustible cigarettes. However, the health burden and effects of their long-term use remain unidentified. This study evaluates the extent and reversibility of cellular damage in human bone marrow-derived mesenchymal stem cells (MSCs), following in vitro exposure to cigarette and e-cigarette smoke aerosols. METHODS:Human primary MSCs were seeded and exposed to combustible or e-cigarette smoke aerosol extracts for 2 or 3 weeks. MSCs were then assessed for survival and membrane protein expression and distribution. In addition, osteogenic differentiation potential was examined after addition of a differentiation cocktail made up of dexamethasone, ascorbic acid, and β-glycerophosphate. RESULTS:MSC-mediated cellular repair, described in terms of MSC proliferation and osteogenic differentiation, was compromised by both cigarette and e-cigarette aerosol extracts. MSCs exhibited partial recovery of their proliferative ability after smoke washout. Their membrane proteins were downregulated, and their distribution was altered on the plasma membrane. MSCs showed signs of early osteogenic differentiation following washout of e-cigarette, but matrix mineralization (indicative of terminal differentiation) was hampered, similarly to combustible cigarette aerosols. CONCLUSIONS:These results provide insights on the safety of e-cigarettes and the potential risks they pose to regenerative health, help explain impaired tissue repair mechanisms in chronic smokers, and can serve to inform future public policies on tobacco control. IMPLICATIONS:This study provides insight on the impact of electronic cigarettes on repair mechanisms mediated by stem cells. Importantly, it also examines the consequences of discontinuing combustible or electronic cigarette smoking on stem cell outcomes, notably proliferation, cell integrity, and differentiation potential. Findings from this work feed into the growing evidence on the harmful and persistent effects of electronic smoking modalities.

Sulfated polysaccharides (SPs) isolated from the marine red macroalga Jania rubens exhibit diverse biological activities. Streptococcus agalactiae (S. agalactiae) poses a particular risk to immunocompromised individuals, including cancer patients. Colorectal cancer (CRC) remains a major cause of cancer-related mortality, and resistance to chemotherapeutics such as doxorubicin (DOX) continues to limit treatment outcomes. This study investigated the antibacterial activity of Lebanese J. rubens SPs against S. agalactiae and evaluated their potential to enhance the efficacy of vancomycin and doxorubicin in human colorectal cancer cell lines (HCT-116, HT-29, Caco-2). Antibacterial activity was assessed using the agar well diffusion method, across extracts, tested at 50–4000 µg/mL. The Minimum Inhibitory Concentration (MIC) was determined and synergistic interactions of SPs and vancomycin were evaluated using the fractional inhibitory concentration index (FICI). Biofilm inhibition assays were conducted, and RT-PCR quantified the expression of genes involved in adhesion and immune evasion. Carbohydrate content was measured using the phenol–sulfuric acid method. Synergistic cytotoxic effects with DOX were evaluated in colorectal cancer cell lines (HCT-116, HT-29, Caco-2) using viability assays, and cytotoxicity was examined in the normal-like colon epithelial cell line NCM460D. SP extracts demonstrated strong antibacterial activity against S. agalactiae, even at lowest concentration tested, with an MIC of 0.39 µg/mL. Synergy with vancomycin was confirmed by FICI values and accompanied by significant inhibition of biofilm formation. The SP–vancomycin combination downregulated key virulence-associated genes, reducing cylE, fbsB2 and CpsG by 74.6

Colorectal cancer (CRC) treatment with standard chemotherapeutics such as capecitabine (CAP) and irinotecan (IRT) is frequently limited by toxicity and resistance. To identify novel adjuvant strategies, we investigated the dichloromethane–methanol (DM) Soxhlet extract of the red alga Jania rubens, previously shown to exert intrinsic antiproliferative effects via reactive oxygen species (ROS) induction, inhibition of epithelial–mesenchymal transition (EMT), and suppression of TET enzymes. The present study evaluated the effect of the DM extract in combination with chemotherapeutic agents in HCT-116, Caco-2, and HT-29 colorectal cancer cell lines, assessing its potential to enhance treatment response. Co-treatment with the DM extract significantly enhanced the cytotoxic and anti-migratory effects of CAP and IRT in HCT-116 and Caco-2 cells. Combination treatment also impaired long-term clonogenic survival, suggesting the inhibition of therapy-resistant subpopulations. Flow cytometric analysis (Annexin V-FITC/PI) revealed a dose-dependent increase in apoptosis in HCT-116 cells following DM treatment. Western blot analysis further supported the pro-apoptotic activity of the DM extract by demonstrating reduced BCL-2 protein expression. The extract additionally modulated the mRNA expression levels of cytokines (TNF-α, IL-6, and IL-10), suggesting potential immunomodulatory effects in colorectal cancer cells. Notably, co-administration of IRT and the DM extract enhanced apoptosis, primarily through the downregulation of the anti-apoptotic gene BCL-2. Together, these findings indicate that the Jania rubens DM extract modulates multiple cellular pathways and enhances the response to conventional CRC chemotherapeutic agents.

Abstract Study purpose: Inflammatory Bowel Disease (IBD) is partly attributed to cytokine storm and infiltrating immune cells. As part of the cell defensive mechanisms, a transcription factor called nuclear factor erythroid 2-related factor 2 (NRF-2) activates genes encoding antioxidant enzymes. Additionally, NRF-2 has recently emerged as an important contributor to chemo-resistance in colon neoplasms, by upregulating multi-drug resistant protein 1 (MDR-1). In addition, epigenetic factors have been shown to influence the onset of IBD and of colorectal cancer. One of these epigenetic factors is the interplay between DNA methylation status. A DNA demethylating enzyme, coded by the ten-to-eleven translocation 2 [tet2], yields the methylcytosine dioxygenase TET-2. Recent studies have demonstrated a potential association between TET-2 and NRF-2 in IBD-associated colon cancer. This study aims to understand the alteration in the expression and activity of these markers under inflammatory conditions. Experimental procedures: NRF-2, MDR-1, and TET-2 expression level was evaluated in parental HT-29 cells and in HT-29 cells downregulated for TET-2 (HT-29shTET-2) and exposed to an inflammatory milieu; at the transcriptional level by quantitative real-time polymerase chain reaction (qPCR). Markers of the epithelial-to-mesenchymal transition (EMT, E-cadherin and N-cadherin) were also evaluated by qPCR and Western blotting. Summary of results: Under inflammatory conditions, NRF-2, MDR-1, and TET-2 expression levels increased. However, NRF-2 and MDR-1 expression levels were downregulated in HT-29shTET-2 cells. Interestingly, N-cadherin levels were higher in HT-29shTET-2 exposed to inflammatory media compared to parental HT-29. Conclusions: NRF-2 and MDR-1 expression levels increase upon exposure of HT-29 cells to inflammation. On the other hand, their expression is downregulated when TET-2 is downregulated, suggesting a potential demethylating role of TET-2 in activating NRF-2 and MDR-1 gene expression. Citation Format: Jessica Saliba, Ali Amhaz, Batoul Moussa, Souraya Ismail, Abdullah Shaito, Marwan El-Sabban. A potential role of NRF-2, MDR-1, and TET-2 in inflammatory bowel disease and colorectal carcinoma: An in vitro study [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2026; Part 1 (Regular Abstracts); 2026 Apr 17-22; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2026;86(7 Suppl):Abstract nr 6577.

The imidazoquinoxaline EAPB0503, an imiquimod analogue, exhibits immunomodulatory and anti-cancer properties against several blood malignancies, including acute myeloid leukemia (AML). The bone marrow (BM) niche plays a pivotal role in normal hematopoiesis and in leukemogenesis. To characterize the molecular potency of EAPB0503 in AML, we compared its effect on the expression profile of genes involved in the function and differentiation potential of BM-derived mesenchymal stem cells (BM-MSCs) from a healthy subject and a patient with AML. Transcriptional levels of stemness markers and genes potentially relevant to niche function were evaluated by quantitative real-time polymerase chain reaction in BM-MSCs isolated from BM aspirates of subjects undergoing BM analysis and exposed to EAPB0503. Differentiation potential was assessed in BM-MSCs exposed to EAPB0503 and induced into osteogenic differentiation using a mixture of dexamethasone, ascorbic acid and β-glycerophosphate (DAG). A colorimetric assay was used to detect early-stage osteogenic differentiation by evaluating alkaline phosphatase (ALP) activity. EAPB0503 treatment induced a differential expression of functionally important marker genes between BM-MSCs from the healthy subject and the patient with AML. As such, levels of Connexin 43 (GJA1) decreased in healthy BM-MSCs and increased in AML-derived BM-MSCs following treatment with EAPB0503. In contrast, transcriptional levels of multi-drug resistance 1 (MDR1 or ABCB1) markedly increased in healthy BM-MSCs and remained stable in AML-derived BM-MSCs. Vascular Endothelial Growth Factor (VEGF) and N-cadherin (CDH2) transcriptional levels increased in AML-derived BM-MSCs, with modest changes in healthy MSCs. In addition, AML-derived BM-MSCs do not seem to respond to differentiation signals in a manner similar to healthy BM-MSCs, as shown by ALP activity. These preliminary findings underscore the differential transcriptional profile in BM-MSCs derived from a healthy subject or a patient with AML. Importantly, a differential gene expression profile was also obtained in response to the therapeutic drug EAPB0503. Our preliminary findings implicate the BM niche in AML leukemogenesis and response to drugs. Souraya Ismail, Fouad Al Feghali, Rana Abdel-Samad, Abdullah A. Shaito, Hiba El Hajj, Carine Deleuze-Masquéfa, Pierre-Antoine Bonnet, Marwan El-Sabban, Jessica Saliba. Differential stemness properties in bone marrow-derived mesenchymal stem cells from healthy subjects and from patients with acute myeloid leukemia [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2025; Part 1 (Regular Abstracts); 2025 Apr 25-30; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2025;85(8_Suppl_1):Abstract nr 16.

Background/Objective: Colon cancer poses a significant health burden, with current treatments often associated with severe side effects and limited effectiveness for some patients. Natural products are gaining interest as adjuvant therapies, potentially reducing side effects and improving responses to conventional treatments. We previously highlighted the potent antineoplastic effects of organic extracts derived from the Lebanese red algae Jania rubens. This study, investigated the anticancer activities of polysaccharide, protein, and lipid extracts from J. rubens, which may serve as adjuvant therapies to enhance conventional treatments. Methods: we employed colorimetric assays, wound healing assays, and cell cycle analysis to evaluate the anticancer activities of the extracts. The polysaccharide extract was characterized for sulfate content and structure using barium chloride-gelatin and FT-IR methods. Results: All J. rubens extracts exhibited significant anticancer effects, with the polysaccharide extract showing particularly strong cytotoxicity, apoptosis induction, and antiproliferative and anti-migratory activities. Conclusion: These findings confirm that J. rubens is a source of bioactive compounds with anticancer potential. Further investigations are needed to elucidate the molecular pathways targeted by J. rubens extracts in cancer cells.

Although 60% of AML patients respond well to standard chemotherapy, most patients eventually relapse, develop chemoresistance, and do not survive more than five years. Targeted therapies, including analogs of imiquimod belonging to the family of imiqualines, emerged as promising agents against AML. Notably, the first-generation imiqualine EAPB0503 proved selective potency against nucleophosmin-1-mutant (NPM1c) AML. Recently, chemical modifications of EAPB0503 led to the development of the lead compound from the second generation, EAPB02303. Here, we demonstrate that EAPB02303 displays 200-fold greater potency, broader activity across AML subtypes, and, importantly, a distinct mechanistic profile when compared to EAPB0503. Unlike EAPB0503, which primarily targeted NPM1c AML cells, EAPB02303 exhibits broad-spectrum activity across various AML subtypes. Remarkably, EAPB02303 anti-leukemic activity was attributed to the inhibition of PI3K/AKT/mTOR signaling activity. Nevertheless, NPM1c AML cells were more sensitive to EAPB02303, likely due to its ability to promote NPM1c protein degradation. In vivo, EAPB02303 potently reduced the leukemic burden and improved organ tumor infiltration in both wt-NPM1 and NPM1c AML xenograft mice. Yet, the significant prolonged survival was exclusive to NPM1c AML xenografts, likely due to superior response conferred by NPM1c degradation. Overall, these findings highlight the potential of EAPB02303 as a powerful therapeutic agent for a range of AML subtypes, supporting its further development for broader clinical use.

While the literature on molecular and clinical effects of smoking on the lungs and other organs has been expansively reviewed, there is no comprehensive compilation of the effects of smoking on stem cell (SC) populations. Recent research has shown that tobacco exposure severely compromises the function of SC populations, particularly those involved in tissue regeneration: mesenchymal SCs (MSCs), neural progenitors, and hematopoietic SCs. SC-based therapies have emerged as a promising approach to counteract smoking-related damage. In particular, MSCs have been extensively studied for their immunomodulatory properties, demonstrating the ability to repair damaged tissues, reduce inflammation, and slow disease progression in conditions such as chronic obstructive pulmonary disease. Combination therapies, which integrate pharmaceuticals with SC treatments, have shown potential in enhancing regenerative outcomes. This review examines the impact of smoking on SC biology, describes the processes impairing SC-mediated repair mechanisms and highlights recent advancements in SC-based therapies in the treatment of smoking-induced diseases. This review has two prongs: (1) it attempts to explain potential smoking-related disease etiology, and (2) it addresses a gap in the literature on SC-mediated repair mechanisms in chronic smokers.

Matrix metalloproteinases (MMPs) are emerging as promising diagnostic and prognostic biomarkers for prostate cancer (PCa). This study explored the association between urinary exosome MMP levels and the incidence of prostate cancer. Urine samples were collected from patients undergoing prostate biopsy or prostatectomy, and from age-matched healthy controls. A total of 147 patients participated, including 37 patients who provided samples before prostatectomy, 41 patients before a biopsy that turned out positive, 21 patients before a biopsy that turned out negative, and 48 healthy controls. The study found that MMP-2 expression was similar in patients with prostate cancer and healthy controls but significantly higher in those with a negative biopsy. MMP-9 expression was elevated in patients with a positive biopsy and even higher in those with a negative biopsy. Multivariate logistic regression, adjusted for age, showed that increased MMP-2 expression was linked to a higher likelihood of a negative biopsy result (OR 1.12 [1.002,1.252], P = 0.046), while increased MMP-9 expression was associated with a higher probability of prostate cancer diagnosis (OR 1.106 [1.002,1.22], P = 0.045). These findings suggest that urinary MMP-9 levels are elevated in PCa patients, though even higher levels in biopsy-negative cases may reflect confounding factors such as benign prostatic inflammation.

Nucleophosmin-1 (NPM1) is a nucleolar chaperone protein frequently mutated in acute myeloid leukemia (AML). ARF and Sentrin/SUMO Specific Peptidase 3 (SENP3) control NPM1 functions through dynamic SUMOylation/de-SUMOylation. Mutated NPM1 is an oncoprotein that exhibits an aberrant cytoplasmic localization (NPM1c) and disrupts PML/P53 signaling. Studies reported increased survival of patients with NPM1c AML when retinoic acid (RA) was added to chemotherapy or hypomethylating agents. Ex vivo, RA initiates NPM1c degradation, P53 activation and cell death. Yet, the molecular mechanisms involved remain elusive. Here we show that in NPM1c AML cell lines or patients’ blasts, NPM1c-triggered mitochondrial dysfunction and oxidative stress drive NPM1c stabilization through SENP3 upregulation. RA decreases mitochondrial ROS production, driving degradation of SENP3, ARF stabilization, PML-dependent NPM1c hyperSUMOylation followed by RNF4-dependent ubiquitination and degradation. Thus, the feedback loop stabilizing NPM1c protein can be interrupted by RA-triggered enhanced mitochondrial fitness, mechanistically explaining the benefit of RA in chemotherapy or hypomethylating agents-treated AMLs.

The red seaweed Jania rubens (J. rubens) is prevalent along the Lebanese coast and has drawn attention for its notable antineoplastic properties. Our previous data showed that its dichloromethane–methanol (DM) extract possesses antioxidant, cytotoxic, and anti-migratory effects on colon cancer cells. In the present study, a GC-MS analysis of DM extract identified a diverse profile of bioactive compounds, including flavonoids and pyrazole derivatives with antioxidant and anticancer activities. In vitro assays demonstrated that the DM extract exerts significant cytotoxic activity against various cancer cell lines, including colon, breast, and cervical types. Further investigation into the underlying molecular mechanisms revealed that the extract induces G2/M cell cycle arrest and reduces the expression of EMT (epithelial–mesenchymal transition) markers, N-cadherin and Twist. In addition, the extract showed anti-metastatic properties through its ability to decrease MMP-2 and MMP-9 activity. Mechanistically, DM caused a substantial reduction in Ten-Eleven Translocation (TET) enzymes TET-1, TET-2, and TET-3, which are essential DNA demethylation regulators, thus decreasing their enzymatic product 5-hydroxymethylcytosine (5-hmC). Interestingly, despite a significant increase in intracellular ROS (reactive oxygen species), suggesting a contribution to cytotoxicity, no substantial change in the biogenesis of promyelocytic leukemia nuclear bodies (PML-NBs) was detected. These findings demonstrate that J. rubens DM extract contains bioactive compounds with multiple anticancer effects, thus making it a promising candidate for developing new therapeutic agents.

Current in vitro models fail to recapitulate specific physiological properties of the human blood-brain barrier (BBB); hence the need for a reliable platform to study central nervous system diseases and drug permeability. To mimic the normally tight blood-brain interface, primary human endothelial cells (HAECs) and primary human astrocytes (A) were grown in a confined space of the physical scaffold created by gelatin methacrylate (GelMA) hydrogel to allow optimal astrocyte-endothelial cell direct/indirect interaction. Evidence for a physiologically relevant BBB was established by assessing the expression of tight junction markers conferring the barrier function, and by measuring biophysical attributes using the trans-endothelial electrical resistance (TEER) and the Evans blue albumin (EBA) permeability assay. An HAEC+A three-dimensional (3D) co-culture was associated with 12-fold higher claudin-5 (CLDN5) and cadherin-1 (CDH1 or Epithelial [E]-cadherin) transcriptional levels than two-dimensional (2D) models. This model conferred the highest TEER (45 Ω·cm2) in 3D HAEC+A, which value was 30 Ω·cm2 in 2D (p < 0.01) and 25 Ω·cm2 in 3D HAEC cultures (p < 0.001). Functionally, in 3D HAEC+A co-cultures, higher TEER resulted in 10-fold and 7-fold lower EBA permeability at 120 min, in HAECs alone or in to 2D co-cultures (p < 0.01). The established human primary cell model has acquired features mimicking the human BBB in vitro, and is now poised to be tested for the permeability of the BBB to pharmacological agents, parasites, cells (such as brain-tropic cancer cell metastasis) and any mechanisms that might involve traversing the BBB.

Toxoplasma gondii is the etiologic agent of toxoplasmosis, a highly prevalent parasitosis. Toxoplasma gondii (T. gondii) transits in the brain from acute (AT) to chronic toxoplasmosis (CT), under host immune control. In immunocompromised patients, reactivation of CT is potentially life-threatening. Behavioral and neurological complications have been associated with CT. Furthermore, an effective treatment targeting CT is still lacking. We previously reported the efficacy of imiquimod against CT. Here, we demonstrate the molecular effects of imiquimod or imiquimod followed by the clinically used combination of sulfadiazine and pyrimethamine (SDZ + PYR) on CT-associated behavior in a rat model. Imiquimod decreased the number of cysts in the brains of chronically infected rats due to an induced reactivation of bradyzoites into tachyzoites. Importantly, this decrease was more pronounced in rats treated with imiquimod followed by SDZ + PYR. Rats chronically infected with T. gondii exhibited an anxiety-like behavior. Notably, treatment with imiquimod reversed this behavior aberrancy, with even a more pronounced effect with imiquimod followed by SDZ/PYR. Similarly, rats chronically infected with T. gondii exhibited learning deficits, and imiquimod alone or followed by SDZ/PYR reversed this behavior. Our results enhance our knowledge of the implications of CT on behavioral aberrancies and highlight the potency of imiquimod followed by SDZ + PYR on these CT-associated complications.

Background/Objectives: A specialized microenvironment in the bone marrow, composed of stromal cells including mesenchymal stem cells (MSCs), supports hematopoietic stem cell (HSC) self-renewal, and differentiation bands play an important role in leukemia development and progression. The reciprocal direct interaction between MSCs and CD34+ HSCs under physiological and pathological conditions is yet to be fully characterized. Methods: Here, we established a direct co-culture model between MSCs and CD34+ HSCs or MSCs and acute myeloid leukemia cells (THP-1, Molm-13, and primary cells from patients) to study heterocellular communication. Results: Following MSCs-CD34+ HSCs co-culture, the expression of adhesion markers N-Cadherin and connexin 43 increased in both cell types, forming gap junction channels. Moreover, the clonogenic potential of CD34+ HSCs was increased. However, direct contact of acute myeloid leukemia cells with MSCs reduced the expression levels of connexin 43 and N-Cadherin in MSCs. The impairment in gap junction formation may potentially be due to a defect in the acute myeloid leukemia-derived MSCs. Interestingly, CD34+ HSCs and acute myeloid leukemia cell lines attenuated MSC osteoblastic differentiation upon prolonged direct cell-cell contact. Conclusions: In conclusion, under physiological conditions, connexin 43 and N-Cadherin interaction preserves stemness of both CD34+ HSCs and MSCs, a process that is compromised in acute myeloid leukemia, pointing to the possible role of gap junctions in modulating stemness.

The interplay between cancer cell physical characteristics and metastatic potential highlights the significance of cancer cell mechanobiology. Using fluidic-based single-cell force spectroscopy (SCFS), quartz crystal microbalance with dissipation (QCM-D), and a model of cells with a spectrum of metastatic potential, we track the progression of biomechanics across the metastatic states by measuring cell-substrate and cell-to-cell adhesion forces, cell spring constant, cell height, and cell viscoelasticity. Compared to highly metastatic cells, cells in the lower spectrum of metastatic ability are found to be systematically stiffer, less viscoelastic, and larger. These mechanical transformations in cells within a cluster correlate with cells' metastatic potential but are significantly absent in single cells. Additionally, the response to chemotherapy is found to be highly dependent on cell viscoelastic properties in terms of both response time and magnitude. Shifts in cell softness and elasticity might serve as mechanoadaptive mechanisms during cancer cell metastasis, contributing to our understanding of metastasis and the effectiveness of potential therapeutic interventions.