Leveraging the 'trans-cleavage′ activity of Cas proteins, the CRISPR/Cas system has successfully transitioned from a gene-editing tool to a new-generation molecular diagnostic platform. This technology demonstrates notable advantages in point-of-care testing of biomarkers, including rapid detection, operational simplicity, and portability. However, it still faces critical challenges such as insufficient sensitivity and limited multiplex detection capability. Recent studies indicate that through innovative strategies including protein engineering, microfluidic integration, and artificial intelligence algorithm optimization, CRISPR/Cas-based diagnostic technology is advancing molecular diagnostics towards field-deployable and intelligent systems, thereby providing novel solutions for intelligent on-site biomarker screening.

Leveraging the 'trans-cleavage′ activity of Cas proteins, the CRISPR/Cas system has successfully transitioned from a gene-editing tool to a new-generation molecular diagnostic platform. This technology demonstrates notable advantages in point-of-care testing of biomarkers, including rapid detection, operational simplicity, and portability. However, it still faces critical challenges such as insufficient sensitivity and limited multiplex detection capability. Recent studies indicate that through innovative strategies including protein engineering, microfluidic integration, and artificial intelligence algorithm optimization, CRISPR/Cas-based diagnostic technology is advancing molecular diagnostics towards field-deployable and intelligent systems, thereby providing novel solutions for intelligent on-site biomarker screening.

Objective To investigate the effects of baicalein (Bai) on acute lung injury (ALI) induced by intestinal ischemia/reperfusion (I/R) and its mechanism in mice.Methods Twenty-four male C57BL/6J mice were divided into three groups by random number table:namely sham group,I/R group and Bai+I/R group,with 8 mice in each group.Intestinal I/R induced lung injury model was reproduced by clamping superior mesenteric artery for 90 minutes,followed by reperfusion.Bai (100 mg/kg) was intraperitoneally injected 1 hour before ischemic challenge in the Bai+I/Rgroup.The mice in sham group underwent the similar procedure with I/R group but without vascular occlusion.All mice were sacrificed at 4 hours of reperfusion,and blood was collected from inferior vena cava and lung tissues were harvested.Lung tissues were stained with hematoxylin-eosin (HE),and histological changes were examined under light microscope for pathological score.Lung wet/dry (W/D) ratio was calculated.Lung cell apoptosis was determined by TdT-mediated dUTP nick end labeling (TUNEL) technique.Serum levels of tumor necrosis factor-α (TNF-α) and interleukin-6(IL-6) were determined by enzyme-linked immunosorbent assay (ELISA).The mRNA expressions of TNF-α and IL-6 in lung tissues were determined by real-time quantitative reverse transcription-polymerase chain reaction (RT-PCR).The protein expression levels of cytoplasmic inhibitory factor-α of nuclear factor-κB (IκB-α) and nucleus NF-κB were determined by Western Blot.Results Under light microscope,a normal lung tissue structure was shown in the sham group and no evidence of obvious lung injury was found.In the I/R group,the alveolar structure was seriously damaged.The alveolar wall was widened and there was significant interstitial edema and leukocytes infiltration.In the Bai+I/R group,pathological damage was significantly decreased as indicated by reduced lung tissue edema and leukocytes infiltration.Compared with the sham group,the lung pathological scores,W/D ratio and cellular apoptosis in the I/R group were significantly increased.Bothserum TNF-α and IL-6 contents and lung TNF-α and IL-6 mRNA expressions were significantly increased.Furthermore,I/R significantly resulted in a decrease of IκB-α in the cytoplasm and an increase of NF-κB in the nucleus.Notably,Bai treatment significantly attenuated ALI induced by intestinal I/R injury.Compared with the I/R group,the lung pathological scores and W/D ratio in the Bai+I/R group were significantly decreased (lung pathological score:4.59±1.17 vs.6.27±1.34,W/D ratio:3.79±0.28 vs.4.32±0.57),cellular apoptosis was significantly decreased [(4.85 ± 2.47)％ vs.(8.15 ± 2.33)％],both serum TNF-α and IL-6 contents and lung TNF-α and IL-6 mRNA expressions were significantly decreased [serum TNF-α (pg/L):124.18±30.49 vs.167.72 ± 38.65,IL-6 (ng/L):1.65 ± 0.69 vs.2.43 ± 0.57;lung TNF-α mRNA (2-△△Ct:4.75 ± 2.38 vs.7.69 ± 2.32,IL-6 mRNA (2-△△ Ct):16.45 ±4.39 vs.27.69 ± 6.82],additionally,Bai pretreatment significantly increased cytoplasmic IκB-α protein expression (gray value:0.47 ± 0.11 vs.0.27 ± 0.09),while decreased nuclear NF-κB protein expression (gray value:0.57 ± 0.13 vs.1.07 ± 0.14,all P ＜ 0.05).Conclusion Bai could attenuate intestinal I/R injury induced ALI via the inhibition of inflammation and apoptosis.

Cytomegalovirus (CMV) is distinct among members of the Herpesviridae family for having the largest dsDNA genome (230 kb). Packaging of large dsDNA genome is known to give rise to a highly pressurized viral capsid, but molecular interactions conducive to the formation of CMV capsid resistant to pressurization have not been described. Here, we report a cryo electron microscopy (cryoEM) structure of the murine cytomegalovirus (MCMV) capsid at a 9.1 Å resolution and describe the molecular interactions among the ∼3000 protein molecules in the MCMV capsid at the secondary structure level. Secondary structural elements are resolved to provide landmarks for correlating with results from sequence-based prediction and for structure-based homology modeling. The major capsid protein (MCP) upper domain (MCPud) contains α-helices and β-sheets conserved with those in MCPud of herpes simplex virus type 1 (HSV-1), with the largest differences identified as a "saddle loop" region, located at the tip of MCPud and involved in interaction with the smallest capsid protein (SCP). Interactions among the bacteriophage HK97-like floor domain of MCP, the middle domain of MCP, the hook and clamp domains of the triplex proteins (hoop and clamp domains of TRI-1 and clamp domain of TRI-2) contribute to the formation of a mature capsid. These results offer a framework for understanding how cytomegalovirus uses various secondary structural elements of its capsid proteins to build a robust capsid for packaging its large dsDNA genome inside and for attaching unique functional tegument proteins outside.
Amino Acid Sequence ; Capsid Proteins ; chemistry ; metabolism ; ultrastructure ; Cryoelectron Microscopy ; Models, Molecular ; Molecular Sequence Data ; Muromegalovirus ; chemistry ; ultrastructure ; Protein Binding ; Protein Multimerization ; Protein Structure, Quaternary ; Protein Structure, Tertiary