Objective To isolate and purify the fusion protein TAT-SOD of superoxide dismutase(SOD) and the protein transduction domain of trans-activator of transcription(TAT) expressed by human immunodeficiency virus typeⅠ(HIV) using the lyophilized powder of recombinant Pichia pastoris fermentation broth as raw material.Methods The pigment and bacterial endotoxin in TAT-SOD were removed by ethanol precipitation, macroporous resin column chromatography, SP-650 ion exchange column chromatography and TritonX-114 phase separation method.Results The enzyme activity recovery rate of 70% ethanol settling TAT-SOD reached(98. 50 ± 1. 71)%, and the purification factor was(1. 04 ± 0. 02). The overall effect of separating and purifying TAT-SOD with macroporous resin was better than SP-650. The purification factor of TATSOD by macroporous resin was(3. 07 ± 0. 06) times, slightly higher than the(2. 78 ± 0. 05) times of SP-650; The removal rate of pigment in TAT-SOD by macroporous resin was(95. 16 ± 1. 65)%, which was much higher than the(44. 73 ± 0. 77)%of SP-650; The enzyme activity recovery rate of TAT-SOD by macroporous resin was(61. 17 ± 1. 06)%, slightly lower than the(69. 93 ± 1. 21)% of SP-650. The purification of TAT-SOD using Triton X-114 phase separation method resulted in a bacterial endotoxin removal rate of(99. 89 ± 1. 73)% and an enzyme activity recovery rate of(95. 02 ± 1. 66)%. The final obtained TAT-SOD had a specific activity of(10 110 ± 175) U/mg.Conclusion The combined method of ethanol precipitation, macroporous resin column chromatography, and TritonX-114 phase separation has good separation and purification effects on TAT-SOD, and is expected to provide a new option for the industrial separation and purification of TAT-SOD.

Ferroptosis of chondrocytes is a significant contributor to osteoarthritis (OA), for which there is still a lack of safe and effective therapeutic drugs targeting ferroptosis. Here, we screen for anti-ferroptotic drugs in Food and Drug Administration (FDA)-approved drug library via a high-throughput manner in chondrocytes. We identified a group of FDA-approved anti-ferroptotic drugs, among which vitamin K showed the most powerful protective effect. Further study demonstrated that vitamin K effectively inhibited ferroptosis and alleviated the extracellular matrix (ECM) degradation in chondrocytes. Intra-articular injection of vitamin K inhibited ferroptosis and alleviated OA phenotype in destabilization of the medial meniscus (DMM) mouse model. Mechanistically, transcriptome sequencing and knockdown experiments revealed that the anti-ferroptotic effects of vitamin K depended on growth arrest-specific 6 (Gas6). Furthermore, exogenous expression of Gas6 was found to inhibit ferroptosis through the AXL receptor tyrosine kinase (AXL)/phosphatidylinositol 3-kinase (PI3K)/AKT serine/threonine kinase (AKT) axis. Together, we demonstrate that vitamin K inhibits ferroptosis and alleviates OA progression via enhancing Gas6 expression and its downstream pathway of AXL/PI3K/AKT axis, indicating vitamin K as well as Gas6 to serve as a potential therapeutic target for OA and other ferroptosis-related diseases.

Objective To investigate whether amplification of?-globin gene can be used to monitor the quality of samples for PCR detection of DNA of Chlamydia trachomatis(CT).Methods First-void urines(FVU),endocervical swabs(ECS),and urethral swabs (UTS) were collected for PCR detection of CT-DNA and cellular?-globin gene.Samples negative for?-globin gene were retested after 10-fold dilution.Results The positive rates of?-globin gene in FVU and ECS were 95.6%(255/264),and 91.7%(413/450) respectively, both higher than that(77.8%,172/221) in UTS(P