Acute respiratory distress syndrome (ARDS) remains one of the most difficult forms of critical lung injury to treat. The syndrome can be triggered by infection, sepsis, trauma or other severe insults, causing the lung’s delicate air–blood barrier to become permeable and flooded with inflammatory fluid. Patients may develop profound oxygen failure, while the repair process itself can leave behind fibrotic tissue that stiffens the lungs. A new study in Molecular Biology Reports describes an experimental cell-therapy strategy designed to redirect macrophages—the immune cells that both amplify inflammation and coordinate tissue repair—toward a more restorative state. Researchers used small interfering RNA, or siRNA, to simultaneously reduce the activity of two genes, Scn1b and Gsdmd, in rat bone-marrow-derived macrophages. When these modified cells were administered to rats with experimentally induced ARDS, they were associated with reduced inflammatory signals in the short term and fewer structural signs of fibrosis one week later.

The approach reflects a growing effort to treat ARDS by changing the behavior of immune cells rather than suppressing the entire immune response. Macrophages are highly adaptable cells that respond to signals in their surroundings. In simplified laboratory terminology, inflammatory macrophages are often described as M1-like, while cells associated with resolution, repair and remodeling are called M2-like. In living tissues, however, macrophages occupy a broad continuum of functional states rather than two fixed categories. The Russian research team first generated bone-marrow-derived macrophages from male Sprague-Dawley rats and characterized them using immunocytochemistry and phagocytosis assays. They then compared gene activity in cells stimulated with lipopolysaccharide, or LPS, to create an inflammatory M1-like state, and cells exposed to interleukin-4 and interleukin-10 to promote an M2-like program. This comparison provided a molecular map for identifying genes that might be manipulated to produce a reparative immune-cell product.

Among the genes that differed between resting and M2-polarized macrophages, Scn1b and Gsdmd emerged as candidates for simultaneous silencing. Scn1b encodes a beta subunit associated with voltage-gated sodium channels. Although sodium-channel components are best known for their roles in excitable cells such as neurons and muscle, ion-channel activity can also influence immune-cell signaling, migration and inflammatory responses. Gsdmd encodes gasdermin D, a key executioner of pyroptosis, an inflammatory form of programmed cell death. When inflammatory caspases cleave gasdermin D, its pore-forming fragment can disrupt the cell membrane and promote the release of cytokines, including interleukin-18. This makes gasdermin D an attractive target when the goal is to limit inflammatory amplification. The researchers used siRNA molecules to reduce expression of both genes at once, producing macrophages designated M(Scn1b+Gsdmd).

The dual knockdown generated a broad change in macrophage biology rather than simply reducing two messenger RNA transcripts. According to the study, gene-expression patterns in the modified cells became more similar to those observed after interleukin-4 and interleukin-10 polarization. Proteomic analyses also indicated changes in cellular pathways, while metabolic measurements showed a shift toward oxidative phosphorylation. This is the process by which mitochondria use nutrients and oxygen to generate ATP, and it is commonly associated with several macrophage programs involved in maintenance and repair, although it is not an exclusive marker of an M2 state. The modified macrophages retained phagocytic activity, an important feature for a potential cell therapy. In contrast, conventionally generated M2 macrophages in the investigators’ experiments displayed significantly reduced phagocytosis. Preserving engulfment capacity could be advantageous in injured lungs, where macrophages must clear cellular debris and dying cells while avoiding excessive inflammatory damage.

The researchers also examined whether substances released by the engineered macrophages could influence other cell types involved in tissue recovery. Conditioned medium collected from M(Scn1b+Gsdmd) cultures improved the proliferation capacity and viability of mesenchymal stem cells, multipotent stromal cells widely investigated for their ability to support tissue repair and regulate inflammation. The study additionally assessed effects on glioblastoma cells, although the central therapeutic interpretation concerned the interaction with stromal and regenerative systems rather than cancer treatment. These experiments suggest that the modified macrophages may act through paracrine communication: soluble proteins, lipids, metabolites or extracellular vesicles released into the local environment could alter the behavior of neighboring cells. The specific factors responsible were not established by the abstract, so the findings should be viewed as evidence of a beneficial secretory influence rather than proof of a single macrophage-derived repair molecule.

The decisive test took place in a rat model of ARDS produced by delivering LPS directly into the trachea. LPS is a component of the outer membrane of many Gram-negative bacteria and activates innate immune receptors, especially Toll-like receptor 4, triggering a cascade that includes nuclear factor kappa B signaling, cytokine release, vascular leakage and recruitment of additional inflammatory cells. This model reproduces selected features of acute inflammatory lung injury but does not capture the full biological diversity of human ARDS, which can arise from multiple causes and may involve ventilation-associated stress, systemic organ failure and infection-specific responses. Following the induction of lung injury, the animals received systemic administration of the siRNA-modified macrophages. Lung and inflammatory outcomes were assessed after 24 hours and again after seven days, allowing the investigators to distinguish early immunomodulation from later tissue-repair effects.

At the 24-hour time point, treatment with M(Scn1b+Gsdmd) was associated with lower levels of IL-18 and TIMP2. Interleukin-18 is a potent inflammatory cytokine that can promote interferon-gamma production and intensify immune activation. Its reduction is consistent with the proposed effect of silencing gasdermin D, although the study does not establish that gasdermin-D-dependent pyroptosis was the sole mechanism responsible. TIMP2, or tissue inhibitor of metalloproteinases 2, regulates matrix metalloproteinases and therefore influences the turnover of extracellular matrix. Altered matrix regulation is central to both wound repair and pathological fibrosis. A decrease in TIMP2 could have complex consequences depending on timing, tissue location and the balance of matrix-degrading enzymes. The researchers’ results indicate that the treatment modified the inflammatory environment early after injury, but further mechanistic work will be needed to determine how these molecular changes connect to macrophage survival, cytokine release and recruitment of host immune cells.

After seven days, the treated animals displayed molecular and histological features interpreted as more reparative. Expression of Vegf, the gene encoding vascular endothelial growth factor, increased in the lungs. VEGF is best known for stimulating blood-vessel growth and vascular permeability, but it also participates in tissue recovery, epithelial responses and communication between endothelial and immune cells. Its role in ARDS is highly context-dependent: excessive VEGF during the acute leak phase can worsen edema, while appropriately timed activity may support repair and restoration of the microvasculature. Histological analysis using Mallory’s trichrome staining showed thinner interalveolar septa and a reduced amount of collagen fibers in animals receiving the modified macrophages. Thickened septa and excess collagen are structural hallmarks of inflammatory remodeling and fibrosis. These observations suggest that the cell treatment may have limited the transition from acute injury to scar-like remodeling, although quantitative lung-function measurements, longer follow-up and comparisons with established treatments would be required to determine whether the anatomical improvements translated into better gas exchange.

The study presents dual gene knockdown as a form of “immune programming,” but its findings remain preclinical and should not be interpreted as evidence of an available ARDS therapy. The experiments used rat cells, male animals and an LPS-driven model, while human ARDS is heterogeneous and often develops in patients with substantial comorbidities. Before clinical translation, researchers would need to examine dosing, cell distribution, persistence, immune compatibility, manufacturing consistency and the possibility of unintended inflammatory or vascular effects. The safety of suppressing gasdermin D also requires careful evaluation because pyroptosis contributes to host defense against some pathogens, and macrophage behavior can vary substantially between tissues. Nevertheless, the work offers a technically distinctive strategy: instead of forcing macrophages into a predetermined state with cytokines, it uses targeted RNA interference to reconfigure an endogenous gene network while preserving phagocytosis. If validated in more clinically representative models, such engineered macrophages could become a platform for moderating early inflammation and reducing the fibrotic consequences of severe lung injury.

Subject of Research: siRNA-engineered macrophage cell therapy for inflammation and fibrosis in acute respiratory distress syndrome

Article Title: Dual knockdown of Scn1b and Gsdmd induces an M2-like functional state in macrophages and modulates inflammation and fibrosis in a rat model of acute respiratory distress syndrome

Article References: Kiseleva V, Vishnyakova P, Kosyreva A, et al. “Dual knockdown of Scn1b and Gsdmd induces an M2-like functional state in macrophages and modulates inflammation and fibrosis in a rat model of acute respiratory distress syndrome.” Molecular Biology Reports 53, 1463 (2026).

Image Credits: AI Generated

DOI: 10.1007/s11033-026-12658-1

Keywords: Acute respiratory distress syndrome (ARDS), macrophages, siRNA, Scn1b, Gsdmd, adoptive cell transfer, cell therapy, inflammation, fibrosis, oxidative phosphorylation

Tags: ARDS treatmentcell-based regenerative therapyfibrosis reduction in lung injurygene knockdown therapyimmune cell reprogramming in critical lung injuryimmune modulation in ARDSM2-like macrophages in lung repairmacrophage polarizationmacrophage-driven inflammationrat model of ARDSScn1b and Gsdmd in lung injurysiRNA therapy for respiratory distress