To evaluate the role of cystatin D as a non-invasive biomarker for primary Sjögren's syndrome (pSS), salivary cystatin D levels were measured and the association between cystatin D and clinical parameters was analysed. A total of 73 patients with pSS, 23 patients with head and neck cancer who had completed radiotherapy (HNCR), and 58 healthy controls (HC) were included in this study. Salivary cystatin D levels were measured via an enzyme-linked immunosorbent assay (ELISA). Salivary gland flow rate, salivary gland ultrasound scores, and disease activity indexes were assessed in patients with pSS. The receiver operating characteristic (ROC) curves was used to assess the potential value of salivary cystatin D as a diagnostic biomarker. Salivary cystatin D levels were significantly reduced in the patients with pSS, compared with the patients with HNCR (p<0.001) and HC (p<0.001). Salivary cystatin D level was positively correlated with unstimulated salivary gland flow rate (USFR) and stimulated salivary gland flow rate (SSFR), whereas, negatively correlated with serum IL-6 levels, IgE levels and peripheral blood CD4+ T cell counts. In addition, salivary cystatin D levels were significantly reduced in the pSS patients with parotid or submandibular gland ultrasonography scores ≥ 2. The diagnostic value of salivary cystatin D was determined by receiver operating curve (ROC) analysis, with an area under the curve of 0.713 (95% CI: 0.632, 0.749, p<0.001), a sensitivity of 51.9% and a specificity of 83.6%. In addition, cystatin D improved the accuracy of pSS diagnosis, particularly in the patient with negative anti-SSA. Salivary cystatin D emerged as a promising biomarker for pSS diagnosis and was correlated with salivary gland dysfunction.

Background: High mobility group box 1 (HMGB1) plays an essential role in various pathological conditions, including inflammation, fibrosis, autoimmune diseases, and carcinogenesis. The quantification of HMGB1 in body fluids holds promise for clinical applications. Objectives: This study aimed to isolate high-affinity single-stranded DNA (ssDNA) aptamers that target HMGB1. Methods: In this study, ssDNA aptamers were selected using Systematic Evolution of Ligands by Exponential Enrichment (SELEX). The affinity and specificity of the aptamers were evaluated through South-Western blot analysis, enzyme-linked aptamer sorbent assay (ELASA), and aptamer-based histochemistry staining. The impact of the aptamers on the biological activity of HMGB1 was tested in the human acute monocytic leukemia cell line, THP-1. Results: An aptamer (H-ap25, dissociation constant = 8.20 ± 0.53 nmol/L) with high affinity for the HMGB1 B box was generated. Further experiments verified that H-ap25 can be used to detect HMGB1 in South-Western blot analysis, ELASA, and aptamer-based histochemistry staining. Moreover, H-ap25 significantly augmented HMGB1-induced expression of tumor necrosis factor-α (TNF-α), interleukin (IL)-1β, IL-6, Toll-like receptor 9 (TLR9), and activation of NF-κB in THP-1 cells. Conclusions: Our results demonstrated that H-ap25 can be used both as an enhancer of HMGB1 and as a probe in research.

Synovial fibroblasts (SFs) transform into an aggressive phenotype in rheumatoid arthritis (RA), playing a significant role in sustaining synovitis and bone erosion. Baricitinib, a JAK inhibitor, has been validated as an effective treatment for RA. However, its direct inhibitory effects on RASFs remain poorly understood. The inflammatory response and aggressive behaviors (proliferation, apoptosis, migration, and invasion) of RASFs were assessed following treatment with varying concentrations of baricitinib. RNA sequencing was performed to identify differentially expressed genes and elucidate the impact of baricitinib on RASFs. The relationship between the JAK-STAT3 signaling pathway and downstream molecules (EGR1, BCL-2, MMP-2, MMP-9) was analyzed via western blotting. Collagen-induced arthritis (CIA) mice were treated with baricitinib via oral gavage or by intra-articular injection of shEGR1 to evaluate therapeutic efficacy. Baricitinib inhibited the production of inflammatory mediators in primary RASFs and reduced the progression of synovial inflammation and hyperplasia in RA patients as assessed by ultrasound. In vitro, baricitinib promoted apoptosis and inhibited the proliferation, migration, and invasion of RASFs, effects associated with decreased expression of the transcriptional regulator EGR1. In vivo, baricitinib effectively reduced arthritis scores, paw swelling, and the expression of inflammatory cytokines in the joint tissues of CIA mice. Similar therapeutic effects were observed upon inhibition of EGR1. Baricitinib inhibits the aggressive phenotype of RASFs through the JAK-STAT3-EGR1 axis, thereby contributing to the alleviation of the inflammatory response and joint destruction in RA.