Biology subjects

Crofford, L. J.

Publications and source records attributed to Crofford, L. J..

5 recordsLinked to original sources

bioRxiv2025

A TLR7/9-IFNα-LDHB axis drives vital NET release and compromises antibacterial defense.

Patients with systemic lupus erythematosus (SLE) are susceptible to bacterial infections, but the underlying dysfunction remains unclear. We found that Staphylococcus aureus triggers mitochondria-dependent suicidal NETosis via lactate sensing in healthy neutrophils, but this response is defective in SLE. Herein, we show that chronic Toll-like receptor (TLR) 7/9 signaling represses mitochondrial lactate dehydrogenase B (LDHB), thereby impairing lactate sensing and downstream suicidal NETosis. Instead, SLE neutrophils default to vital NET release; a less bactericidal, type I interferon (IFN)-driven process amplified by staphylococcal pore-forming toxins and sustained by elevated systemic IFN levels observed in SLE. Combined treatment with hydroxychloroquine (HCQ) and interferon-alpha/beta receptor (IFNAR) blockade restores LDHB expression, NET homeostasis, and bacterial clearance in lupus-prone mice. Neutrophils from SLE patients exhibit similar defects, which are reversed by HCQ and the IFNAR-blocking antibody anifrolumab. These findings identify a clinically actionable immunometabolic checkpoint linking chronic autoimmune signaling to defective antibacterial defense in SLE.

immunology↗

bioRxiv2023

Low Gut Microbial Diversity Augments Estrogen-driven Pulmonary Fibrosis in Female-Predominant Interstitial Lung Disease

Although profibrotic cytokines such as IL-17A and TGF-{beta}1 have been implicated in interstitial lung disease (ILD) pathogenesis, interactions between gut dysbiosis, gonadotrophic hormones and molecular mediators of profibrotic cytokine expression, such as phosphorylation of STAT3, have not been defined. Here we show by chromatin immunoprecipitation sequencing (ChIP-seq) analysis of primary human CD4+ T cells that regions within the STAT3 locus are significantly enriched for binding by the transcription factor estrogen receptor alpha (ERa). Using the murine model of bleomycin-induced pulmonary fibrosis, we found significantly increased regulatory T cells compared to Th17 cells in the female lung. Genetic absence of ESR1 or ovariectomy in mice significantly increased pSTAT3 and IL-17A expression in pulmonary CD4+ T cells, which was reduced after repletion of female hormones. Remarkably, there was no significant reduction in lung fibrosis under either condition, suggesting that factors outside of ovarian hormones also contribute. Assessment of lung fibrosis among menstruating females in different rearing environments revealed that environments favoring gut dysbiosis augment fibrosis. Furthermore, hormone repletion following ovariectomy further augmented lung fibrosis, suggesting pathologic interactions between gonadal hormones and gut microbiota on lung fibrosis severity. Analysis in female sarcoidosis patients revealed a significant reduction in pSTAT3 and IL-17A levels and a concomitant increase in TGF-{beta}1 levels in CD4+ T cells, compared to male sarcoidosis patients. These studies reveal that estrogen is profibrotic in females and that gut dysbiosis in menstruating females augments lung fibrosis severity, supporting a critical interaction between gonadal hormones and gut flora in lung fibrosis pathogenesis.

immunology↗

bioRxiv2021

Translating outcomes from the clinical setting to preclinical models: chronic pain and functionality in chronic musculoskeletal pain

Fibromyalgia (FM) is a chronic pain disorder characterized by chronic widespread musculoskeletal pain (CWP), tenderness, and fatigue, which interferes with daily functioning and quality of life. In clinical studies, this symptomology is assessed, while preclinical models of CWP are limited to nociceptive assays. The aim of the study was to investigate the human-to-model translatability of clinical behavioral assessments for pain and muscle function in a preclinical model of CWP. We assessed correlations between pain behaviors and muscle function in a preclinical model of CWP and in women with fibromyalgia to examine whether similar relationships between outcomes existed in both settings, for usability of clinical assays in model systems. For preclinical measures, the acidic saline model of FM which induces widespread muscle pain, was used in adult female mice. Two gastrocnemius injections of acidic or physiological pH saline were given following baseline measures, five days apart. An array of adapted pain measures and functional assays were assessed for three weeks. For clinical measures, pain and functional assays were assessed in adult women with FM. For both preclinical and clinical outcomes, movement-evoked pain (MEP) was associated with mechanical pain sensitivity. Mechanical sensitivity was correlated to shifts in weight-bearing preclinically and was predictive of functionality in patients. Preclinically, it is imperative to expand how the field assesses pain behaviors when studying multi- symptom disorders like FM. Targeted pain assessments to match those performed clinically is an important aspect of improving preclinical to clinical translatability of animal models. SummaryPreclinical assessments of chronic musculoskeletal pain recapitulate several outcome measures for clinical assessment of patients with FM, particularly prolonged resting pain, and MEP.

neuroscience↗

bioRxiv2020

Matrix-targeted Nanoparticles for MMP13 RNA Interference Blocks Post-Traumatic Osteoarthritis.

Osteoarthritis (OA) is a debilitating and prevalent chronic disease, but there are no approved disease modifying OA drugs (DMOADs), only pharmaceuticals for pain management. OA progression, particularly for post-traumatic osteoarthritis (PTOA), is associated with inflammation and enzymatic degradation of the extracellular matrix. In particular, Matrix Metalloproteinase 13 (MMP13) breaks down collagen type 2 (CII), a key structural component of cartilage extracellular matrix, and consequently, matrix degradation fragments perpetuate inflammation and a degenerative cycle that leads to progressive joint pathology. Here, we tested targeted delivery of endosome-escaping, MMP13 RNA interference (RNAi) nanoparticles (NPs) as a DMOAD. The new targeting approach pursued here deviates from the convention of targeting specific cell types (e.g., through cell surface receptors) and instead leverages a monoclonal antibody (mAbCII) that targets extracellular CII that becomes uniquely accessible at early OA focal defects. Targeted mAbCII-siNPs create an in situ NP depot for retention and potent activity within OA joints. The mAbCII-siNPs loaded with MMP13 siRNA (mAbCII-siNP/siMMP13) potently suppressed MMP13 expression (95% silencing) in TNF-stimulated chondrocytes in vitro, and the targeted mAbCII-siNPs had higher binding to trypsin-damaged porcine cartilage than untargeted control NPs. In an acute mechanical injury mouse model of PTOA, mAbCII-siNP/siMMP13 achieved 80% reduction in MMP13 expression (p = 0.00231), whereas a non-targeted control achieved only 55% silencing. In a more severe, PTOA model, weekly mAbCII-siNP/siMMP13 long-term treatment provided significant protection of cartilage integrity (0.45+/- .3 vs 1.6+/-.5 on the OARSI scale; p=0.0166), and overall joint structure (1.3+/-.6 vs 2.8+/-.2 on the Degenerative Joint Disease scale; p<0.05). Intra-articular mAbCII-siNPs better protected articular cartilage (OARSI score) relative to either single or weekly treatment with the clinical gold stand steroid treatment methylprednisolone. Finally, multiplexed gene expression analysis of 254 inflammation-related genes showed that MMP13 inhibition suppressed clusters of genes associated with tissue restructuring, angiogenesis (associated with synovial inflammation and thickening), innate immune response, and proteolysis. This work establishes the new concept of targeting unique local extracellular matrix signatures to sustain retention and increase delivery efficacy of biologics with intracellular activity and also validates the promise of MMP13 RNAi as a DMOAD in a clinically-relevant therapeutic context. O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=148 SRC="FIGDIR/small/925321v1_ufig1.gif" ALT="Figure 1"> View larger version (54K): org.highwire.dtl.DTLVardef@1803795org.highwire.dtl.DTLVardef@c1254dorg.highwire.dtl.DTLVardef@78233eorg.highwire.dtl.DTLVardef@151d96f_HPS_FORMAT_FIGEXP M_FIG O_FLOATNOAbstract Figure:C_FLOATNO PTOA targeted delivery of MMP13 siRNA to block disease progressionThe top left schematic illustrates the progression (left to right) from healthy knee joint, through inflammation induction following traumatic injury, to cartilage loss and degenerative joint disease (including synovial response). Degradation of cartilage enhances inflammation, inducing a degenerative cycle (middle right). The bottom of the graphic illustrates the concept of the matrix targeted nanocarriers for enhanced retention and activity of MMP13 siRNA at sites of cartilage injury. C_FIG

bioengineering↗

bioRxiv2018

Nonsteroidal anti-inflammatory drugs alter the microbiota and exacerbate Clostridium difficile colitis while dysregulating the inflammatory response

Clostridium difficile infection (CDI) is a major public health threat worldwide. The use of nonsteroidal anti-inflammatory drugs (NSAIDs) is associated with enhanced susceptibility to and severity of nosocomial CDI; however, the mechanisms driving this phenomenon have not been elucidated. NSAIDs alter prostaglandin (PG) metabolism by inhibiting cyclooxygenase (COX) enzymes. Here, we found that treatment with the NSAID indomethacin prior to infection altered the microbiota and dramatically increased mortality and intestinal pathology associated with CDI in mice. We demonstrate that in C. difficile-infected animals, indomethacin lead to PG deregulation, an altered proinflammatory transcriptional and protein profile, and perturbed epithelial cell junctions. These effects were paralleled by an increased recruitment of intestinal neutrophils and CD4+ cells. Together, these data implicate NSAIDs in perturbation of the gut microbiota and disruption of protective COX-mediated PG production during CDI, resulting in altered epithelial integrity and associated immune responses.

microbiology↗