IL-1β–Induced Inflammatory Transcriptome and TGF-β1 Attenuation of Chemokine Expression in ACL Remnant Fibroblasts

Background: The high incidence of posttraumatic osteoarthritis after anterior cruciate ligament (ACL) injury and reconstruction suggests that biological mechanisms beyond joint instability contribute to disease progression. After injury, ACL-resident cells are exposed to inflammatory mediators within the joint and may influence the local inflammatory microenvironment; however, their global transcriptomic responses to inflammatory stimuli remain underexplored. Hypothesis: Interleukin-1 beta (IL-1β) induces inflammatory transcriptomic responses in ACL remnant–derived (ACLr) fibroblasts, and transforming growth factor beta 1 (TGF-β1) attenuates these responses. Study Design: Descriptive laboratory study. Methods: Primary ACLr fibroblasts were isolated from ACL remnants (n = 10) collected at the time of knee surgery. Near-confluent cultures were treated with IL-1β (10 ng/mL) for 24 hours to model an inflammatory joint environment. Genome-wide transcriptional changes were assessed by RNA sequencing using an Illumina NovaSeq-6000, with selected targets confirmed by microfluidic digital polymerase chain reaction (PCR). To evaluate the effects of TGF-β1, ACLr fibroblasts pretreated with IL-1β were exposed to TGF-β1 (10 ng/mL) for 48 hours, and chemokine transcript levels were assessed using digital PCR. Results: IL-1β induced broad inflammatory transcriptional responses characterized by upregulation of chemokines ( CXCL6 , CXCL8 , CXCL5 , CCL20 , CXCL1 , and CXCL3 ), cytokines ( IL33 , IL1B , and IL6 ), and matrix-degrading enzymes ( MMP3 and MMP12 ), while suppressing extracellular matrix-associated genes ( KRT14 , COL21A1 , POSTN , COL1A1 , and COL6A3 ). Functional enrichment analysis demonstrated activation of immune and inflammatory responses, cytokine and chemokine signaling, and cell activation. TGF-β1 reduced the expression of all assessed IL-1β–induced chemokines, with reduction of up to 50%, with statistically significant suppression observed for 13 of the 16 chemokines. Concomitantly, TGF-β1 treatment reduced expression of CEBPB , NFKBIA , and NFKBIZ , transcripts implicated in inflammatory gene regulation. Conclusion: ACLr fibroblasts mounted a robust inflammatory transcriptomic response to IL-1β, characterized by marked induction of chemokines and inflammatory pathways. TGF-β1 attenuated many of these responses, supporting its role as a regulator of inflammatory signaling in ACL-resident cells. Clinical Relevance: ACL-resident cells respond to injury and may contribute to the post-injury inflammatory microenvironment. Therapeutic modulation of inflammatory signaling in these cells could represent a strategy to mitigate early molecular events associated with post-traumatic osteoarthritis following ACL tear.