Osteoarthritis (OA) causes chronic pain that significantly impairs quality of life, with current treatments often proving insufficient and accompanied by adverse effects. Recent research has identified the dorsal root ganglion (DRG) and its resident macrophages as crucial mediators of chronic OA pain through neuroinflammation driven by macrophage polarization. We present a novel injectable thermo-sensitive hydrogel system, KAF@PLEL, designed to deliver an anti-inflammatory peptide (KAF) specifically to the DRG. This biodegradable hydrogel enables sustained KAF release, promoting the reprogramming of DRG macrophages from pro-inflammatory to anti-inflammatory phenotypes. Through comprehensive in vitro and in vivo studies, we evaluated the hydrogel's biocompatibility, effects on macrophage polarization, and therapeutic efficacy in chronic OA pain management. The system demonstrated significant capabilities in preserving macrophage mitochondrial function, suppressing neuroinflammation, alleviating chronic OA pain, reducing cartilage degradation, and improving motor function in OA rat models. The sustained-release properties of KAF@PLEL enabled prolonged therapeutic effects while minimizing systemic exposure and side effects. These findings suggest that KAF@PLEL represents a promising therapeutic approach for improving outcomes in OA patients through targeted, sustained treatment.

BACKGROUND: The autophagy of macrophage cells plays a key role in regulating the immune system and inflammatory response, while knockdown Atg5 gene can specifically inhibit the autophagy. There are some shortcomings of traditional siRNA transfection methods such as transience and incompleteness. It is of great value and significance to construct a stable Atg5 gene knockdown cell line in RAW 264.7 cells for studying the macrophage autophagy and immune inflammation-related diseases and their pathogenesis. OBJECTIVE: To construct RAW 264.7 macrophage cell line stably knocking down the Atg5 gene by lentivirus infection, and to provide chassis cells for studying macrophage autophagy and immune inflammation-related diseases and their pathogenesis. METHODS: Recombinant expression vector (HBLV-m-Atg5-shRNA-GFP-Puro) with green fluorescence signal, Puro resistance gene, and knockdown Atg5 gene was constructed and transfected into 293T cells to obtain lentivirus plasmid system. The green fluorescence signal of the acquired lentivirus infected RAW 264.7 cells was observed under an inverted fluorescence microscope. Purinomycin resistance screening and flow cytometry were used to obtain high-purity infected cells. The RAW 264.7 cell line stably knocking down the Atg5 gene was identified by real-time quantitative polymerase chain reaction and western blot assay. RESULTS AND CONCLUSION: DNA sequencing results showed that Atg5 gene interfering sequences were correctly inserted into the expression vector, and the HBLV-m-Atg5-shRNA-GFP-Puro expression vector was successfully constructed. After infecting RAW 264.7 cell line with lentivirus plasmid, green fluorescence was observed under an inverted fluorescence microscope. Green fluorescent protein positive cell groups were observed by flow cytometry. The results of real-time quantitative polymerase chain reaction and western blot assay showed that the expression level of the Atg5 gene in RAW 264.7 cell line was significantly decreased (P < 0.05), indicating that the RAW 264.7 cell line with stable knockdown of Atg5 gene was successfully constructed.

ObjectiveTo investigate the distribution of high-intensity zone (HIZ) of lumbar intervertebral disc in patients with low back and/or leg pain,and analyze its related factors.MethodsSix hundred and twenty-eight patients with low back and/or leg pain were examined by MRI scan from June 2009 to August 2010.According to the diagnostic criteria of HIZ,the features of distribution of HIZ on age,segment and degree of intervertebral disc degeneration were analyzed retrospectively.ResultsAmong 3140 intervertebral discs of the 628 patients,172 cases (27.39％,172/628) and 206 discs (6.56％,206/3140)were involved with HIZ.There was no significant difference between men and women [26.38％(86/326)vs.28.48％(86/302)] (P=0.556).HIZ occurred more often [40.22％(72/179)] in those patients between 40 and 49 years of age.The incidence of HIZ at the segments from L1-2,L2-3,L3-4,L4-5,L5-S1 was 0.80％(5/628),2.07％(13/628),2.07％(13/628),14.01％(88/628) and 13.85％(87/628) respectively.In cases with and without HIZ,the incidence of intervertebral disc degeneration up to grade V was 49.03％(101/206) and 23.76％(697/2934) respectively (P ＜ 0.01 ).HIZ was correlated with age,degree of intervertebral disc degeneration and disc segment (r =-0.040,P=0.025 ;r =0.217,P＜ 0.01 ;r =0.179,P＜ 0.01 ).Conclusions HIZ is correlated with age,degree of intervertebral disc degeneration and disc segment.Intervertebral disc degeneration plays the most important role in the occurence of HIZ.HIZ_mainly occur in L4-5 and L5-S1 segment and in those between 40 and 49 years of age.