Traditional development of small protein scaffolds has relied on display technologies and mutation-based engineering, which limit sequence and functional diversity, thereby constraining their therapeutic and application potential. Protein design tools have significantly advanced the creation of novel protein sequences, structures, and functions. However, further improvements in design strategies are still needed to more efficiently optimize the functional performance of protein-based drugs and enhance their druggability. Here, we extended an evolution-based design protocol to create a novel minibinder, BindHer, against the human epidermal growth factor receptor 2 (HER2). It not only exhibits super stability and binding selectivity but also demonstrates remarkable properties in tissue specificity. Radiolabeling experiments with ^99mTc, ⁶⁸Ga, and ¹⁸F revealed that BindHer efficiently targets tumors in HER2-positive breast cancer mouse models, with minimal nonspecific liver absorption, outperforming scaffolds designed through traditional engineering. These findings highlight a new rational approach to automated protein design, offering significant potential for large-scale applications in therapeutic mini-protein development.

AIM:This study aims to investigate the sensitizing effects of lycorine(Lyc)in combination with cisplatin(Cis)on human osteosarcoma cells and to explore the underlying mechanisms of action.METHODS:Human osteosarcoma cell lines MG63,HOS,and cisplatin-resistant HOS/DDP cells were utilized to evaluate the effects of Lyc and cisplatin,both alone and in combination,on cell viability using the CCK8 assay.The clonogenic assay was performed to assess cell proliferation capacity,while the scratch assay evaluated the drugs' effects on cell migration.Reactive oxygen species(ROS)levels were measured using a ROS assay kit,and changes in intracellular glutathione(GSH)levels were assessed with a GSH/oxidized glutathione(GSSG)assay kit.The mitochondrial membrane potential was analyzed via JC-1 staining to determine the drugs' effects on mitochondrial function.Intracellular iron(Ⅱ)content changes were detected us-ing FerrOrange,a fluorescence probe,and cellular malondialdehyde(MDA)levels were measured using an MDA assay kit.RT-qPCR was employed to evaluate the expression levels of key genes related to ferroptosis,and Western blot analysis was conducted to detect changes in the protein expression levels of Yes-associated protein 1(YAP1),acyl-CoA synthetase long-chain family member 4(ACSL4),glutathione peroxidase 4(GPX4),heme oxygenase-1(HO-1),and transferrin re-ceptor(TFRC).RESULTS:Both Lyc and cisplatin effectively inhibited the proliferation of human osteosarcoma cells.Notably,the combination of Lyc and cisplatin led to a more substantial reduction in cell viability,proliferation,and migra-tion abilities in MG63,HOS,and HOS/DDP cells compared to cisplatin alone.Additionally,this combination significant-ly increased ROS levels while decreasing GSH content,indicating mitochondrial damage and elevated iron(Ⅱ)and MDA levels.RT-qPCR results revealed that the combination treatment more significantly downregulated ferroptosis-promoting genes and upregulated ferroptosis-inhibiting genes compared to cisplatin treatment alone(P<0.05).Western blot results showed a slight decrease in GPX4 protein expression following Lyc and cisplatin treatment,while expression levels of YAP1,TFRC,ACSL4,and HO-1 were significantly increased(P<0.05).CONCLUSION:Lyc enhances the sensitivi-ty of MG63,HOS,and HOS/DDP cells to cisplatin by promoting ferroptosis through the YAP1/TFRC signaling pathway.

Objective:To investigate the anti-inflammatory activity of alcohol extract of Polyrhachis dives(PDAEs)against rheumatoid arthritis(RA)in vitro and in vivo.Methods:In vivo and in vitro anti-RA inflammatory response of PDAEs was investigated using a rat model of collagenous arthritis induced by bovine type Ⅱ collagen and an LPS-induced RAW264.7 cell inflammatory model.Results:PDAEs inhibited the polarization of M1 type macrophages in vivo and in vitro,reduced expressions of TNF-α,IL-1β,IL-6,iNOS,and promoted polarization of M2 type macrophages,enhanced expressions of anti-inflammatory cytokines,such as IL-10 and TGF-β,so as to achieve the anti-inflammatory effect.The experiments in vivo also showed that PDAEs had the immunomodulatory effect,the potential mechanism may be the regulation of Th17/Treg balance by regulating the expression of PD-1 and TGF-β,thus correcting the over-strong autoimmune response.Conclusion:PDAEs may reduce the inflammatory reaction of RA through anti-inflam-matory and immunomodulatory effects.

Objective:To investigate the anti-inflammatory activity of alcohol extract of Polyrhachis dives(PDAEs)against rheumatoid arthritis(RA)in vitro and in vivo.Methods:In vivo and in vitro anti-RA inflammatory response of PDAEs was investigated using a rat model of collagenous arthritis induced by bovine type Ⅱ collagen and an LPS-induced RAW264.7 cell inflammatory model.Results:PDAEs inhibited the polarization of M1 type macrophages in vivo and in vitro,reduced expressions of TNF-α,IL-1β,IL-6,iNOS,and promoted polarization of M2 type macrophages,enhanced expressions of anti-inflammatory cytokines,such as IL-10 and TGF-β,so as to achieve the anti-inflammatory effect.The experiments in vivo also showed that PDAEs had the immunomodulatory effect,the potential mechanism may be the regulation of Th17/Treg balance by regulating the expression of PD-1 and TGF-β,thus correcting the over-strong autoimmune response.Conclusion:PDAEs may reduce the inflammatory reaction of RA through anti-inflam-matory and immunomodulatory effects.

AIM:This study aims to investigate the sensitizing effects of lycorine(Lyc)in combination with cisplatin(Cis)on human osteosarcoma cells and to explore the underlying mechanisms of action.METHODS:Human osteosarcoma cell lines MG63,HOS,and cisplatin-resistant HOS/DDP cells were utilized to evaluate the effects of Lyc and cisplatin,both alone and in combination,on cell viability using the CCK8 assay.The clonogenic assay was performed to assess cell proliferation capacity,while the scratch assay evaluated the drugs' effects on cell migration.Reactive oxygen species(ROS)levels were measured using a ROS assay kit,and changes in intracellular glutathione(GSH)levels were assessed with a GSH/oxidized glutathione(GSSG)assay kit.The mitochondrial membrane potential was analyzed via JC-1 staining to determine the drugs' effects on mitochondrial function.Intracellular iron(Ⅱ)content changes were detected us-ing FerrOrange,a fluorescence probe,and cellular malondialdehyde(MDA)levels were measured using an MDA assay kit.RT-qPCR was employed to evaluate the expression levels of key genes related to ferroptosis,and Western blot analysis was conducted to detect changes in the protein expression levels of Yes-associated protein 1(YAP1),acyl-CoA synthetase long-chain family member 4(ACSL4),glutathione peroxidase 4(GPX4),heme oxygenase-1(HO-1),and transferrin re-ceptor(TFRC).RESULTS:Both Lyc and cisplatin effectively inhibited the proliferation of human osteosarcoma cells.Notably,the combination of Lyc and cisplatin led to a more substantial reduction in cell viability,proliferation,and migra-tion abilities in MG63,HOS,and HOS/DDP cells compared to cisplatin alone.Additionally,this combination significant-ly increased ROS levels while decreasing GSH content,indicating mitochondrial damage and elevated iron(Ⅱ)and MDA levels.RT-qPCR results revealed that the combination treatment more significantly downregulated ferroptosis-promoting genes and upregulated ferroptosis-inhibiting genes compared to cisplatin treatment alone(P<0.05).Western blot results showed a slight decrease in GPX4 protein expression following Lyc and cisplatin treatment,while expression levels of YAP1,TFRC,ACSL4,and HO-1 were significantly increased(P<0.05).CONCLUSION:Lyc enhances the sensitivi-ty of MG63,HOS,and HOS/DDP cells to cisplatin by promoting ferroptosis through the YAP1/TFRC signaling pathway.

【Objective】 To evaluate the screening efficacy and practical value of the portable microfluidic biochemical analyzer in the detection of blood donors before blood donation. 【Methods】 Blood donor samples, clinical blood samples and constant quality control products were collected. Referring to the documents of ISO15189 and National Health Industry Standard, the precision and accuracy of hemoglobin (Hb) and alanine aminotransferase (ALT) were verified and compared with other detection systems. 【Results】 The MS200 biochemistry instrument has an intra-batch precision of 1.40% to 1.46%, inter-batch precision of 1.91% to 1.94%, and correctness bias of -0.9% to -1.3% for Hb test, and an intra-batch precision of 3.77% to 4.86%, inter-batch precision of 4.92% to 6.02%, and correctness bias of -3.0% to -4.8% for ALT test, which were within the range of quality requirements of industry standard. Comparison of Hb test results between MS200 biochemistry and Hb201 analyser on 1 189 peripheral blood samples from donors showed no statistically significant difference (P>0.05). 65 samples showed positive correlation between MS200 biochemistry and XS-900i automated hematology analyzer on Hb test results (R²=0.986, P=0.000). Correlation analysis of all the results of ALT detection by MS200 biochemical analyzer and AU480 biochemical analyzer in 1 065 samples showed a positive correlation (R²=0.965, P=0.000). The elevated ALT samples did not affect the Hb test results, and the samples with abnormal Hb value did not affect the ALT test results, with no interference between the two items in the detection. 【Conclusion】 The MS200 biochemical analyzer based on microfluidic technology has reliable methodological performance and can meet the need of pre-donation testing.

Background: The incidence density of metabolic dysfunction-associated fatty liver disease (MAFLD) and the effect of a healthy lifestyle on the risk of MAFLD remain unknown. We evaluated the prevalence and incidence density of MAFLD and investigated the association between healthy lifestyle and the risk of MAFLD. Methods: A cross-sectional analysis was conducted on 37,422 participants to explore the prevalence of MAFLD. A cohort analysis of 18,964 individuals was conducted to identify the incidence of MAFLD, as well as the association between healthy lifestyle and MAFLD. Cox proportional hazards regression was used to calculate the hazard ratio (HR) and 95% confidence interval (CI) with adjustments for confounding factors. Results: The prevalence of MAFLD, non-alcoholic fatty liver disease, and their comorbidities were 30.38%, 28.09%, and 26.13%, respectively. After approximately 70 thousand person-years of follow-up, the incidence densities of the three conditions were 61.03, 55.49, and 51.64 per 1,000 person-years, respectively. Adherence to an overall healthy lifestyle was associated with a 19% decreased risk of MAFLD (HR, 0.81; 95% CI, 0.72 to 0.92), and the effects were modified by baseline age, sex, and body mass index (BMI). Subgroup analyses revealed that younger participants, men, and those with a lower BMI experienced more significant beneficial effects from healthy lifestyle. Conclusion Our results highlight the beneficial effect of adherence to a healthy lifestyle on the prevention of MAFLD. Health management for improving dietary intake, physical activity, and smoking and drinking habits are critical to improving MAFLD.

Objective:To establish the UPLC fingerprint analysis method for Puerariae Lobamle Radix. Methods:The column was Agilent ZORBAX Eclipse Plus C18 column (2.1 mm×100 mm, 1.8 μm). The mobile phase consisted of acetonitrile (A)-0.1% formic acid (b), gradient elution, flow rate was 0.3 ml/min. The detection wavelength was 250 nm. The column temperature was set at 30 ℃. The sample volume was 2 μl, and the similarity evaluation system of traditional Chinese medicine fingerprint was adopted. The cluster analysis, principal component analysis and partial least square method in SPSS software were used to judge the differences of Pueraria lobata from different habitats.Results:With puerarin as reference peak, 22 common peaks were calibrated, and 6 peaks were identified. The similarity of 25 batches of samples was above 0.990 except S22 was 0.935, which indicated that the samples were with good consistency. Through cluster analysis, principal component analysis and partial least square analysis, 25 batches of Puerariae Lobamle Radix can be clustered into 4-5 categories, and different components such as 3'-hydroxy puerarin were found. The extraction process of Puerariae Lobamle Radix was optimized based on analytic hierarchy process and multi-index orthogonal test. The results showed that adding 50% ethanol 40 ml and refluxing for 40 min was the best extraction process. Conclusion:UPLC fingerprint is suitable for Puerariae Lobamle Radix, and the results are reliable. It can be used as a quality evaluation method for Puerariae Lobamle Radix.

Objective:To investigate the safety and efficacy of Tubridge flow diverter for the treatment of recurrent internal carotid blood blister-like aneurysms after stent-assisted embolization.Methods:From June 2018 to April 2021, patients with recurrent internal carotid blood blister-like aneurysms treated with Tubridge flow diverter in the Department of Neurosurgery, Changhai Hospital, Naval Medical University were enrolled retrospectively. The perioperative safety, immediate postoperative and follow-up results were analyzed.Results:A total of 6 patients with recurrent internal carotid blood blister-like aneurysm after stent-assisted embolization were enrolled. The time interval from the first stent-assisted embolization to Tubridge placement was 14 to 90 d. Tubridge implantation alone was used in 4 patients, and Tubridge was implanted in the other 2 patients after the coils were packed. There were no complications during the perioperative period, and no rebleeding was observed after clinical follow-up for 5 to 36 months. Five patients were followed up by angiography for 1-3 months, and the aneurysms disappeared completely.Conclusion:Tubridge flow diverter for the treatment of recurrent internal carotid blood blister-like aneurysms is safe and effective.

In addition to causing high disability and high fatality rates, ruptured intracranial aneurysms can also cause cognitive impairment. Although preventive surgical treatment can avoid intracranial aneurysm rupture and bleeding, patients may still have a certain degree of cognitive impairment, even in patients with good clinical recovery after surgery. There is no systematic review on the effect of different surgical methods on cognitive function, and the best surgical method is still inconclusive. This article reviews the cognitive impairment in patients with intracranial aneurysm, hoping to provide a basis for clinical treatment decisions.