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 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 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 observe the clinical efficacy of Yin-Yang balancing acupuncture plus Tuina(Chinese therapeutic massage)for post-stroke depression(PSD). Methods:A total of 72 PSD patients were randomized into an acupuncture-Tuina group and a Western medicine group using the random number table method,with 36 cases in each group.The Western medicine group was treated with fluoxetine hydrochloride capsules.The acupuncture-Tuina group was treated with Yin-Yang balancing acupuncture plus Tuina therapy.Both groups were treated for 4 weeks.After treatment,the clinical efficacy was compared,and the changes in Hamilton depression scale(HAMD)score,National Institute of Health stroke scale(NIHSS)score,serum neurotransmitter levels,and electroencephalogram(EEG)were observed. Results:The total effective rate of the Western medicine group and the acupuncture-Tuina group was 83.3%and 94.4%,respectively.There was a significant difference in the total effective rate between the two groups(P<0.05).After treatment,the serum levels of 5-hydroxytryptamine(5-HT),norepinephrine(NE),and dopamine(DA)were higher than those in the same group before treatment,and the intra-group differences were all statistically significant(P<0.05).The serum levels of 5-HT,NE,and DA in the acupuncture-Tuina group were higher than those in the Western medicine group,and the differences between the groups were statistically significant(P<0.05).After treatment,the scores of HAMD and NIHSS in both groups decreased,and the intra-group differences were statistically significant(P<0.05).The scores in the acupuncture-Tuina group were lower than those in the Western medicine group,and the differences between the two groups were all statistically significant(P<0.05).After treatment,the EEG diffuse wave[(delta+theta)/(alpha+beta)]ratio(DTABR)and delta/beta ratio(DBR)in both groups decreased,and the intra-group differences were statistically significant(P<0.05).The ratios in the acupuncture-Tuina group were lower than those in the Western medicine group,and the differences between the two groups were statistically significant(P<0.05). Conclusion:Yin-Yang balancing acupuncture plus Tuina therapy has a better efficacy than oral fluoxetine hydrochloride capsules in the treatment of PSD,which can increase the levels of serum 5-HT,DA,and NE,and regulate brain waves.

Objective To investigate the effects of combined exercise training on exercise ability and inflammatory factors in elderly patients with chronic obstructive pulmonary disease(COPD)complicated with sarcopenia.Methods Sixty elderly patients with COPD complicated with sarcopenia who were treated in Hangzhou Third People's Hospital from July 2022 to May 2023 were selected as study objects,and were divided into control group and treatment group according to random number table method,with 30 cases in each group.The control group received routine treatment of COPD,while the treatment group received combined exercise training on the basis of routine treatment for 12 weeks.The appendicular skeletal muscle mass index(ASMI),grip strength,walking speed,6 min walking distance(6MWD),COPD assessment test(CAT),forced expiratory volume in one second(FEV1)/forced vital capacity(FVC)、serum interleukin-6(IL-6)and tumor necrosis factor-α(TNF-α)were compared.The correlation between serum IL-6、TNF-α and ASMI,grip strength、walking speed were analyzed.Results After treatment,6MWD in both groups was significantly longer than before treatment,and CAT score was significantly lower than before treatment(P＜0.05).The ASMI,grip strength and walking speed of patients in treatment group were significantly higher than before treatment,and the serum IL-6 and TNF-α were significantly lower than before treatment(P＜0.05).The ASMI,grip strength and walking speed of treatment group were significantly higher than those of control group,6MWD was significantly longer than that of control group,CAT score,serum IL-6 and TNF-α were significantly lower than those of control group(P＜0.05).There was no significant difference in FEV1/FVC between two groups before and after treatment(P＞0.05).Pearson correlation analysis showed that serum IL-6、TNF-α were negatively correlated with ASMI、grip strength and walking speed.Conclusion Combined exercise training can improve muscle mass and strength,improve exercise ability and clinical symptoms,and inhibit inflammatory response in elderly patients with COPD complicated with sarcopenia.

Objective To explore the clinical efficacy of Fuling Zexie Decoction combined with lifestyle intervention in patients with metabolic syndrome (MetS) of dampness syndrome. Methods A total of 40 patients with MetS of dampness syndrome were randomly divided into experimental group (n=20) and control group (n=20) according to a randomized double-blind placebo-controlled design. The experimental group was given oral administration of Fuling Zexie Decoction,the control group was given placebo,150 mL each time,twice a day. Both groups were given healthy lifestyle(healthy eating habits and exercise mode) education. The intervention period was 16 weeks. Traditional Chinese medicine dampness syndrome scale score and the diagnostic indexes of MetS,including waist circumference (WC),systolic blood pressure (SBP),diastolic blood pressure (DBP),triglyceride (TG),high density lipoprotein cholesterol (HDL-C) and fasting blood glucose (FBG),were evaluated before and after treatment. Other metabolic indexes such as body mass index(BMI),total cholesterol(TC),low density lipoprotein cholesterol(LDL-C),homeostasis model assessment of insulin resistance(HOMA-IR),uric acid(UC),as well as inflammation related indexes including C-reactive protein (CRP),interleukin-6 (IL-6) and tumor necrosis factor(TNF-α) were also evaluated. Results A total of 33 people completed the trial,including 17 people in the experimental group and 16 people in the control group. Compared to the baseline period,FBG,BMI,LDL-C and dampness syndrome scale score in the experimental group were significantly decreased at the 8th and 16th week of intervention(P<0.05). WC was significantly decreased at the 8th week of intervention(P<0.05). CRP showed a downward trend at the 16th week of intervention(P<0.05). Moreover,SBP and IL-6 in control group decreased significantly at the 8th and 16th week of intervention(P<0.05). WC,DBP and HOMA-IR decreased significantly at the 8th week of intervention (P<0.05). There was no significant difference in other outcome indexes between the two groups at the 8th and 16th week of intervention (P>0.05). The safety indexes of all patients before and after intervention were not significantly abnormal,and no serious adverse events occurred. Conclusion This study initially shows the positive regulatory effect of Fuling Zexie Decoction combined with lifestyle on metabolism and inflammation-related indexes in people with MetS of dampness syndrome,which helps to improve the level of dampness syndrome and provides a basis for further research.

Objective To observe the value of breast imaging reporting and data system(BI-RADS)classification combined with mammography(MG)radiomics for differentiating breast benign and malignant amorphous calcification lesions.Methods A total of 217 breast amorphous calcification lesions in 206 female patients,including 43 malignant and 174 benign ones were retrospectively enrolled and randomly divided into training set(n=151)and validation set(n=66)at the ratio of 7∶3.Then radiomics features were screened based on breast MG images.BI-RADS model,radiomics model and combined model were established,and the value of each model for distinguishing benign and malignant lesions was observed.Results Eight optimal radiomics features were selected.The area under the curve(AUC)of BI-RADS model,radiomics model and combined model for differentiating breast benign and malignant amorphous calcification lesions was 0.821,0.763 and 0.897 in training set,0.800,0.746 and 0.893 in validation set,respectively,AUC of combined model was significantly higher than that of the other two(both P＜0.05).Conclusion BI-RADS classification combined with MG radiomics was helpful for differential diagnosis of breast benign and malignant amorphous calcification lesions,with efficacy higher than that of each alone.

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.