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.

Hypoxic-ischemic (HI) brain damage poses a high risk of death or lifelong disability, yet effective treatments remain elusive. Here, we demonstrated that miR-100-5p levels in the lesioned cortex increased after HI insult in neonatal mice. Knockdown of miR-100-5p expression in the brain attenuated brain injury and promoted functional recovery, through inhibiting the cleaved-caspase-3 level, microglia activation, and the release of proinflammation cytokines following HI injury. Engineered extracellular vesicles (EVs) containing neuron-targeting rabies virus glycoprotein (RVG) and miR-100-5p antagonists (RVG-EVs-Antagomir) selectively targeted brain lesions and reduced miR-100-5p levels after intranasal delivery. Both pre- and post-HI administration showed therapeutic benefits. Mechanistically, we identified protein phosphatase 3 catalytic subunit alpha (Ppp3ca) as a novel candidate target gene of miR-100-5p, inhibiting c-Fos expression and neuronal apoptosis following HI insult. In conclusion, our non-invasive method using engineered EVs to deliver miR-100-5p antagomirs to the brain significantly improves functional recovery after HI injury by targeting Ppp3ca to suppress neuronal apoptosis.
Animals ; MicroRNAs/metabolism* ; Extracellular Vesicles/metabolism* ; Mice ; Recovery of Function/physiology* ; Hypoxia-Ischemia, Brain/therapy* ; Mice, Inbred C57BL ; Antagomirs/administration & dosage* ; Male ; Animals, Newborn ; Apoptosis/drug effects* ; Brain Injuries/metabolism* ; Glycoproteins ; Peptide Fragments ; Viral Proteins

Disruption of normal secretion or mucociliary clearance can impair airway defense mechanisms and lung function, and increase the risk of infection. Airway clearance techniques are recommended as part of a comprehensive treatment plan for patients. Among these, vibratory expectoration is an important method of airway clearance, which loosens and liquefies mucus and metabolites on the surface of the respiratory tract through chest wall oscillation, promoting ciliary movement to facilitate sputum expulsion. However, commonly used handheld vibrating head devices and vest-type vibration expectorators have several limitations in clinical practice, such as inconvenience of operation, limited treatment time, poor adaptability, and difficulty in disinfection. To address these issues, the research team from the department of critical care medicine at Ruijin Hospital, Shanghai Jiao Tong University, has designed a novel belt-type vibration expectorator, which has been granted a national utility model patent (Patent No.: ZL 2023 2 1610983.1). The device is mainly composed of a chest strap assembly, a sputum clearance component, and a fixed shoulder strap component. Several pockets are placed on the outer surface of the chest strap, with corresponding inner-side openings that allow the percussion head of the percussive expectorator placed inside the pocket to make contact with the patient's chest wall. Each pocket has markings indicating the percussion position, enabling the placement of the percussive expectorator according to the location of infection, thereby achieving multi-point, precise percussive vibration expectoration in different body positions. On the inner side of the chest strap, there are diagrams illustrating postural drainage, providing guidance on the body positions patients should assume based on the location of infection. The hook-and-loop fasteners on both sides of the chest strap can be wrapped around and secured according to the patient's body shape, ensuring that the sputum clearance components adhere tightly to the chest wall, allowing the vibrations generated by percussion to be effectively transmitted to the patient's airways. Additionally, to prevent the chest strap from slipping due to changes in the patient's position, a Y-shaped fixing strap can be selectively attached to the chest strap for further stabilization. This innovation not only simplifies the operation process, improves convenience and flexibility of use, but also supports the principle of "disinfection after each use by one person," which helps to reduce the risk of nosocomial infections and improve the efficiency of patients' respiratory rehabilitation.
Humans ; Vibration ; Equipment Design ; Chest Wall Oscillation/instrumentation* ; Sputum ; Expectorants ; Mucociliary Clearance

Objective To investigate the application effect of postural gravity traction combined with floss and titanium clip pulley external traction in endoscopic submucosal dissection(ESD)for early intestinal lesions.Methods A total of 100 patients with early colorectal lesions admitted to the Affiliated Hospital of Jiangsu University from January 2022 to September 2024 were selected as the research subjects and divided in-to the observation group and the control group,with 50 cases in each group.The control group underwent con-ventional intestinal ESD treatment,while the observation group used positional gravity traction combined with dental floss and titanium clips to form pulley external traction in ESD treatment.Clinical data including opera-tion time,number of submucosal injections,intraoperative blood loss,lesion resection effect,complication inci-dence,and hospital stay were compared between the two groups.Results The total operation time in the ob-servation group was shorter than that in the control group,and the total number of submucosal injections was less than that in the control group,with statistically significant differences(P<0.05).There were no signifi-cant differences in intraoperative blood loss,complete resection rate,complication incidence,en bloc resection rate,and hospital stay between the two groups(P>0.05).For lesions≤1 cm or>5 cm in size,there were no significant differences in operation time,complete resection rate and en bloc resection rate between the two groups(P>0.05).For lesions>1-3 cm or>3-5 cm in size and laterally spreading lesions,significant differences were observed in operation time,number of submucosal injections,complete resection rate,and en bloc resection rate between the two groups(P<0.05).For pedunculated polyps,there were no significant differences in the number of submucosal injections,complete resection rate and en bloc resection rate between the two groups(P>0.05),but the operation time differed significantly(P<0.05).Conclusion Postural gravity traction combined with dental floss and titanium clip to form pulley external traction is simple to oper-ate in ESD for early intestinal lesions.It can maintain a clear field of view,shorten operation time,reduce the incidence of complications,and is safe and effective.

Objective To investigate the dietary patterns of rural residents in the high-incidence areas of esophageal cancer (EC), and to explore the clustering and influencing factors of risk factors associated with high-incidence characteristics. Methods A special structured questionnaire was applied to conduct a face-to-face survey on the dietary patterns of rural residents in Yanting county of Sichuan Province from July to August 2021. Univariate and multivariate logistic regression models were used to analyze the influencing factors of risk factor clustering for EC. Results There were 838 valid questionnaires in this study. A total of 90.8% of rural residents used clean water such as tap water. In the past one year, the people who ate fruits and vegetables, soybean products, onions and garlic in high frequency accounted for 69.5%, 32.8% and 74.5%, respectively; the people who ate kimchi, pickled vegetables, sauerkraut, barbecue, hot food and mildew food in low frequency accounted for 59.2%, 79.6%, 68.2%, 90.3%, 80.9% and 90.3%, respectively. The clustering of risk factors for EC was found in 73.3% of residents, and the aggregation of two risk factors was the most common mode (28.2%), among which tumor history and preserved food was the main clustering pattern (4.6%). The logistic regression model revealed that the gender, age, marital status and occupation were independent influencing factors for the risk factors clustering of EC (P<0.05). Conclusion A majority of rural residents in high-incidence areas of EC in Yanting county have good eating habits, but the clustering of some risk factors is still at a high level. Gender, age, marital status, and occupation are influencing factors of the risk factors clustering of EC.

Objective To improve the efficiency of induced differentiation of primitive neural epithelial cells derived from human induced pluripotent stem cells(hiPSCs-NECs)into functional midbrain dopaminergic progenitor cells(DAPs).Methods HiPSCs were cultured in mTeSRTM medium containing DMH1(10 μmol/L),SB431542(10 μmol/L),SHH(200 ng/mL),FGF8(100 ng/mL),purmorphamine(2 μmol/L),CHIR99021(3 μmol/L),and N2(1%)for 12 days to induce their differentiation into primitive neuroepithelial cells(NECs).The hiPSCs-NECs were digested with collagenase IV and then cultured in neurobasal medium supplemented with 1%N2,2%B27-A,BDNF(10 ng/mL),GDNF(10 ng/mL),AA,TGF-β,cAMP,and 1%GlutaMax in the presence of different concentrations of Rho kinase inhibitor Y27632,and the culture medium was changed the next day to remove Y27632.Continuous induction was performed until day 28 to obtain DAPs.Results Human iPSCs expressed the pluripotency markers OCT4,SOX2,Nanog,and SSEA1 and were positive for alkaline phosphatase staining.The hiPSCs-NECs were obtained on day 13 in the form of neural rosettes expressing neuroepithelial markers SOX2,nestin,and PAX6.In digested hiPSCs-NECs,the addition of 5 μmol/L Y27632 significantly promoted survival of the adherent cells,increased cell viability and the proportion of S-phase cells(P<0.01),and reduced the rate of apoptotic cells(P<0.05).On day 28 of induction,the obtained cells highly expressed the specific markers of DAPS(TH,FOXA2,NURR1,and Tuj1).Conclusion Treatment with Y27632(5 μmol/L)for 24 h significantly promotes the survival of human iPSCs-NECs during their differentiation into DPAs without affecting the cell differentiation,which indirectly enhances the efficiency of cell differentiation.

Objective:To evaluate the efficacy of calaglitzin combined with Degu insulin Liraglutide and pioglitazone metformin in the treatment of obese type 2 diabetes mellitus (T2DM) .Methods:Ninety-nine obese T2DM patients admitted to our hospital from Apr. 2021 to Jun. 2023 were divided into 2 groups according to different treatment methods. The control group ( n=52) were treated with Delgu insulin liraglutide and pioglitazone metformin; the observation group ( n=47) were treated with cagaglizin on the basis of the control group, and both groups were treated for 8 weeks. The therapeutic effect of the two groups was compared. Results:The effective rate of observation group was 95.74% (45/47), higher than that of control group 82.69% (43/52) ( P<0.05). After eight weeks of treatment, the levels of FPG (7.96±0.74) mmol/L, 2hPBG (8.36±1.02) mmol/L and BMI (24.63±1.51) kg/m 2 in the observation group were higher than those in the control group ( P<0.05). After 8 weeks of treatment, HOMA-IR (2.41±0.56) of observation group was lower than that of control group (2.84±0.61), GIR (5.63±0.97) mg·kg -1·min -1 was higher than that of the control group (5.17±0.95) mg·kg -1·min -1 ( P<0.05). After eight weeks of treatment, the levels of TG (4.22±0.43) mmol/L, TC (2.15±0.21) mmol/L and ApoB (1.18±0.14) mmol/L in the observation group were higher than those in the control group (2.49±0.26) mmol/L, (4.69±0.47) mmol/L and (1.45±0.19) mmo l/L ( P<0.05). The incidence of adverse reactions in the observation group was 8.51% (4/47), lower than that in the control group (5.77% (3/52) ( P>0.05) . Conclusion:Taking calaglitzin in combination with Delgu insulin liraglutide and pioglitazone metformin can significantly reduce insulin resistance, increase the sensitivity of peripheral tissues to insulin, reduce blood glucose level and BMI level, and the drug safety is reasonable.

Objective To improve the efficiency of induced differentiation of primitive neural epithelial cells derived from human induced pluripotent stem cells(hiPSCs-NECs)into functional midbrain dopaminergic progenitor cells(DAPs).Methods HiPSCs were cultured in mTeSRTM medium containing DMH1(10 μmol/L),SB431542(10 μmol/L),SHH(200 ng/mL),FGF8(100 ng/mL),purmorphamine(2 μmol/L),CHIR99021(3 μmol/L),and N2(1%)for 12 days to induce their differentiation into primitive neuroepithelial cells(NECs).The hiPSCs-NECs were digested with collagenase IV and then cultured in neurobasal medium supplemented with 1%N2,2%B27-A,BDNF(10 ng/mL),GDNF(10 ng/mL),AA,TGF-β,cAMP,and 1%GlutaMax in the presence of different concentrations of Rho kinase inhibitor Y27632,and the culture medium was changed the next day to remove Y27632.Continuous induction was performed until day 28 to obtain DAPs.Results Human iPSCs expressed the pluripotency markers OCT4,SOX2,Nanog,and SSEA1 and were positive for alkaline phosphatase staining.The hiPSCs-NECs were obtained on day 13 in the form of neural rosettes expressing neuroepithelial markers SOX2,nestin,and PAX6.In digested hiPSCs-NECs,the addition of 5 μmol/L Y27632 significantly promoted survival of the adherent cells,increased cell viability and the proportion of S-phase cells(P<0.01),and reduced the rate of apoptotic cells(P<0.05).On day 28 of induction,the obtained cells highly expressed the specific markers of DAPS(TH,FOXA2,NURR1,and Tuj1).Conclusion Treatment with Y27632(5 μmol/L)for 24 h significantly promotes the survival of human iPSCs-NECs during their differentiation into DPAs without affecting the cell differentiation,which indirectly enhances the efficiency of cell differentiation.

Objective To investigate the risk factors affecting the malignancy of amorphous calcifications in the breast and to establish a predictive nomogram.Methods Patients with amorphous calcifications detected by mammography were retrospectively collected,clinical data were obtained from electronic medical record(EMR),and the mammographic features of the patients were assessed by diagnostic physicians.The risk factors affecting the malignancy of amorphous calcifications were analyzed to develop a predictive model and to assess the performance of the model.Results A total of 153 amorphous calcifications in 144 patients were included in the study,and the overall malignancy rate of calcifications was 20.92％.Patient's age ≥45 years,linear distribution of calcifications,unilateral single or unilateral multiple calcifications,and a larger maximum ratio of calcification extent all predicted a higher probability of malignancy,establishing a nomogram based on these 4 risk factors,with a 3.65％predicted probability of malig-nancy as the cut-off,33.99％(52/153)of patients were allowed to be spared biopsy.Conclusion Patient's age and the distribution,number,and maximum ratio of calcifications may be the risk predictors of malignancy for amorphous calcifications,with nomogram con-struction for distinguishing benignity from malignancy of amorphous calcifications via combining with mammographic features and clinical data.

The clinical efficacy advantages of traditional Chinese medicine （TCM） compound prescriptions have always been inadequately characterized in experimental research，which has become a bottleneck restricting the development of TCM pharmacology and even the progress of TCM. The concept of simultaneous treatment/regulation，guided by the theory of mutual generation and restriction of five zang-organs，has guiding significance in the clinical practice of TCM throughout history and is still widely used in the current clinical practice. However，this unique and clinically valuable diagnostic and therapeutic medication system based on the syndrome differentiation has been completely ignored in the modern research of TCM pharmacology，which might be one of the key factors restricting the pharmacological characterization of the therapeutic advantages of TCM compound prescriptions. On the basis of systematically summarizing the phased progress and achievements of the efficacy evaluation of TCM compound prescriptions，this article explores the path of exploring the pharmacological advantages of TCM compound prescriptions on simultaneous treatment/regulation on the basis of the correlation patterns of five zang-organs，from the theory of Zangxiang，the core concept of five zang-organs，and the TCM disease recognition based on the theory of mutual generation and restriction of five zang-organs. With the heart-lung correlation as a breakthrough point，this study explored a new characterization method for the pharmacological advantages of TCM，aiming to provide new ideas for evaluating the efficacy of TCM compound prescriptions.