In recent years, the application of artificial intelligence (AI) in the field of biology has witnessed remarkable advancements. Among these, the most notable achievements have emerged in the domain of protein structure prediction and design, with AlphaFold and related innovations earning the 2024 Nobel Prize in Chemistry. These breakthroughs have transformed our ability to understand protein folding and molecular interactions, marking a pivotal milestone in computational biology. Looking ahead, it is foreseeable that the accurate prediction of various physicochemical properties of proteins—beyond static structure—will become the next critical frontier in this rapidly evolving field. One of the most important protein properties is thermodynamic stability, which refers to a protein’s ability to maintain its native conformation under physiological or stress conditions. Accurate prediction of protein stability, especially upon single-point mutations, plays a vital role in numerous scientific and industrial domains. These include understanding the molecular basis of disease, rational drug design, development of therapeutic proteins, design of more robust industrial enzymes, and engineering of biosensors. Consequently, the ability to reliably forecast the stability changes caused by mutations has broad and transformative implications across biomedical and biotechnological applications. Historically, protein stability was assessed via experimental methods such as differential scanning calorimetry (DSC) and circular dichroism (CD), which, while precise, are time-consuming and resource-intensive. This prompted the development of computational approaches, including empirical energy functions and physics-based simulations. However, these traditional models often fall short in capturing the complex, high-dimensional nature of protein conformational landscapes and mutational effects. Recent advances in machine learning (ML) have significantly improved predictive performance in this area. Early ML models used handcrafted features derived from sequence and structure, whereas modern deep learning models leverage massive datasets and learn representations directly from data. Deep neural networks (DNNs), graph neural networks (GNNs), and attention-based architectures such as transformers have shown particular promise. GNNs, in particular, excel at modeling spatial and topological relationships in molecular structures, making them well-suited for protein modeling tasks. Furthermore, attention mechanisms enable models to dynamically weigh the contribution of specific residues or regions, capturing long-range interactions and allosteric effects. Nevertheless, several key challenges remain. These include the imbalance and scarcity of high-quality experimental datasets, particularly for rare or functionally significant mutations, which can lead to biased or overfitted models. Additionally, the inherently dynamic nature of proteins—their conformational flexibility and context-dependent behavior—is difficult to encode in static structural representations. Current models often rely on a single structure or average conformation, which may overlook important aspects of stability modulation. Efforts are ongoing to incorporate multi-conformational ensembles, molecular dynamics simulations, and physics-informed learning frameworks into predictive models. This paper presents a comprehensive review of the evolution of protein thermodynamic stability prediction techniques, with emphasis on the recent progress enabled by machine learning. It highlights representative datasets, modeling strategies, evaluation benchmarks, and the integration of structural and biochemical features. The aim is to provide researchers with a structured and up-to-date reference, guiding the development of more robust, generalizable, and interpretable models for predicting protein stability changes upon mutation. As the field moves forward, the synergy between data-driven AI methods and domain-specific biological knowledge will be key to unlocking deeper understanding and broader applications of protein engineering.

Objective:To investigate the prognostic value of minimal residual disease(MRD)detected by multi-parameter flow cytometry(MFC)in pediatric patients with acute myeloid leukemia(AML)after induction chemotherapy.Methods:A retrospective study was conducted on 97 pediatric patients initially diagnosed with AML at Wuhan Children's Hospital from August 2015 to December 2022.The study analyzed the results of MRD detection using MFC after the first and second cycles of induction chemotherapy,and its association with prognosis were analyzed.Results:Following the first cycle of induction treatment,57 of the 97 patients tested positive for MRD(MRD1+,58.8％).Subsequently,19 patients remained MRD positive(MRD2+,19.6％)after the second cycle of induction treatment.Kaplan-Meier survival analysis showed that the estimated 3-year overall survival(OS)rate of the 37(64.9％)MRD1+patients who underwent transplantation was significantly higher than that of the 20(35.1％)MRD1+patients who did not undergo transplantation(84.6％vs 40.0％,P=0.0001).Among the 35 MRD1+MRD2-patients,the 3-year OS rate of the 25 children who underwent transplantation was higher than that of the 10 children who did not undergo transplantation(87.2％vs 70.0％,P=0.3229).The 3-year OS rate of the 19 MRD1+MRD2+patients was lower than that of the 35 MRD1+MRD2-patients(57.4％vs 81.8％,P=0.059).In the 19 MRD2+patients,the 3-year OS rate of the 12 children who underwent transplantation was significantly higher than that of the 7 children who did not undergo transplantation(80.8％vs 14.3％,P=0.0007).There was no significant difference in 3-year OS between the 12 MRD1+MRD2+patients and 25 MRD1+MRD2-patients,both treated with transplantation(80.8％vs 87.2％,P=0.8868).In those not treated with transplantation,the 7 MRD1+MRD2+patients had a significantly lower 3-year OS compared with the 10 MRD1+MRD2-patients(14.3％vs 70.7％,P=0.0114).Further multivariate analysis indicated that MRD2 positivity and transplantation were both independent prognostic factors(P=0.031,0.000),while MRD1 positivity was not significantly associated with the overall prognosis of 97 patients(P=0.902).Conclusion:MRD positivity following the second cycle of induction chemotherapy is an independent risk factor for unfavorable outcomes in children with AML.MRD2 positivity indicates a poorer prognosis and can help to identify the candidates requiring transplantation.MRD2 positivity is not a contraindication for transplantation in pediatric patients,and early transplantation significantly improves the prognosis of high-risk patients.

Objective:To evaluate the clinical characteristics and risk factors of infections occurring during hospitalization in patients with multiple myeloma (MM) treated with new generation therapies (including immuno-modulatory drugs,proteasome inhibitors and monoclonal antibodies).Methods:The clinical data were collected from 155 patients with multiple myeloma who were treated in Shanxi Bethune Hospital from March,2017 to March,2022 and were retrospectively analyzed.For this study,the following therapies were considered to be new generation therapies:lenalidomide,pomadomide,bortezomib,ixazomib,daratumumab. The clinical characteristics and risk factors of infection were analyzed.Results:A total of 155 patients were included in this study.The median follow-up time was 20 months.Of 155 patients with MM,242 infection episodes were identified.Among the 242 infections,the incidence of clinically defined infection (CDI)was the highest (186,76.86％),followed by microbiologically defined infection (MDI)in 50 cases (20.66％),and fever at unknown focus (FUF)in 6 cases (2.48％).35 cases (14.46％)of bacteria,10 cases (4.13％)of viruses,and 5 cases (2. 07％)of fungi were clearly infected.The most common site of infection was the lower respiratory tract in 90 cases (37.19％),the upper respiratory tract in 83 cases (34.30％),and the digestive tract in 27 cases (11.16％).All-cause mortality was 8.39％(13/155).In univariate analysis,there was a higher correlation between ISS stage Ⅲ,the number of treatment lines ≥2,frail and infected patients with multiple myeloma.In multivariate analysis,ISS stage Ⅲ(OR=2.96,95％CI:1.19-7.40,P=0.02),the number of treatment lines ≥2 (OR=2.91,95％CI:1.13-7.51,P=0.03)and frail (OR=3.58,95％CI:1.44-8.89,P=0. 01)were risk factors for infection in patients with multiple myeloma in the era of new drugs.Conclusion:Patients with multiple myeloma treated with new agents are prone to bacterial infection during hospitalization.ISS stage Ⅲ,lines of therapy(≥2)and frail were associated with high risk for infection.

Objective:To compare the therapeutic effects of 0.05% cyclosporine and 0.1% fluorometholone eye drops in patients with moderate and severe dry eye.Methods:A randomized controlled study was conducted.Fifty-two patients (52 eyes) with moderate to severe dry eye in Tianjin Medical University Eye Hospital from August 2021 to December 2022 were enrolled and randomly divided into 0.05% cyclosporine group and 0.1% fluorometholone group by random number table method, with 26 cases (26 eyes) in each group.Patients received 0.05% cyclosporine eye drops (2 times/day) and 0.1% fluorometholone eye drops (2 times/day) combined with calf blood deproteinized extract eye drops (4 times/day) according to the grouping.Before and 1, 3 and 6 months after treatment, clinical symptoms and signs were observed and Ocular Surface Disease Index (OSDI) score, corneal fluorescein staining (CFS) score, Schirmer Ⅰ test (SⅠT), non-invasive first tear film break-up time (NIBUTf), and conjunctival goblet cell (CGC) density were recorded.Before treatment and after 6 months of treatment, changes in corneal nerves and dendritic cells (DC) were observed by in vivo confocal microscopy (IVCM).This study adhered to the Declaration of Helsinki and was approved by the Medical Ethics Committee of Eye Hospital of Tianjin Medical University (No.2021KY-17).Written informed consent was obtained from each subject. Results:Compared with the 0.1% fluorometholone group, CFS score decreased after 1 month of treatment, but SⅠT, NIBUTf and CFS score increased after 3 months of treatment, and OSDI score, SⅠT and CFS score decreased after 6 months of treatment in the 0.05% cyclosporine group, showing statistically significant differences (all at P<0.05).Compared with baseline, in the 0.05% cyclosporine group, NIBUTf increased and CFS score decreased after 1 month of treatment, OSDI score and CFS score decreased, SⅠT and NIBUTf increased after 3 and 6 months of treatment, showing statistically significant differences (all at P<0.05).In the 0.1% fluorometholone group, CFS score decreased after 3 months of treatment, OSDI score and CFS score decreased, SⅠT increased after 6 months of treatment compared to baseline, showing statistically significant differences (all at P<0.05).OSDI score and CFS score decreased, SⅠT increased after 6 months of treatment compared to 3 months of treatment in the 0.05% cyclosporine group, and the differences were statistically significant (all at P<0.05).Baseline and CGC densities after 1, 3 and 6 months of treatment were (147.66±17.29), (195.44±15.46), (210.36±19.15) and (282.09±22.63)cells/mm 2 in the 0.05% cyclosporine group and (138.09±17.29), (95.67±15.46), (117.77±19.15) and (109.13±22.63)cells/mm 2 in the 0.1% fluorometholone group, respectively, with a statistically significant overall difference ( Fgroup=11.724, P<0.001; Ftime=4.837, P=0.005).Compared with the 0.1% fluorometholone group, CGC density in the 0.05% cyclosporine group increased after 1, 3 and 6 months of treatment, with statistically significant differences (all at P<0.05).Compared with baseline, the CGC density increased in the 0.05% cyclosporine group after 1, 3 and 6 months of treatment, and the differences were statistically significant (all at P<0.05).Compared with the 0.1% fluorometholone group, the corneal nerve fiber density in the 0.05% cyclosporine group increased after 6 months of treatment, and corneal DC density, area and dendrite number decreased, showing statistically significant differences (all at P<0.05). Conclusions:Cyclosporine 0.05% eye drops combined with calf blood deproteinized extract eye drops can improve symptoms and signs in patients with moderate to severe dry eye, and the long-term effect is better than that of 0.1% fluorometholone plus calf blood deproteinized extract eye drops.

Objective To investigate the incidence rate,clinical characteristics,and prognosis of neonatal stroke in Shenzhen,China.Methods Led by Shenzhen Children's Hospital,the Shenzhen Neonatal Data Collaboration Network organized 21 institutions to collect 36 cases of neonatal stroke from January 2020 to December 2022.The incidence,clinical characteristics,treatment,and prognosis of neonatal stroke in Shenzhen were analyzed.Results The incidence rate of neonatal stroke in 21 hospitals from 2020 to 2022 was 1/15 137,1/6 060,and 1/7 704,respectively.Ischemic stroke accounted for 75％(27/36);boys accounted for 64％(23/36).Among the 36 neonates,31(86％)had disease onset within 3 days after birth,and 19(53％)had convulsion as the initial presentation.Cerebral MRI showed that 22 neonates(61％)had left cerebral infarction and 13(36％)had basal ganglia infarction.Magnetic resonance angiography was performed for 12 neonates,among whom 9(75％)had involvement of the middle cerebral artery.Electroencephalography was performed for 29 neonates,with sharp waves in 21 neonates(72％)and seizures in 10 neonates(34％).Symptomatic/supportive treatment varied across different hospitals.Neonatal Behavioral Neurological Assessment was performed for 12 neonates(33％,12/36),with a mean score of(32±4)points.The prognosis of 27 neonates was followed up to around 12 months of age,with 44％(12/27)of the neonates having a good prognosis.Conclusions Ischemic stroke is the main type of neonatal stroke,often with convulsions as the initial presentation,involvement of the middle cerebral artery,sharp waves on electroencephalography,and a relatively low neurodevelopment score.Symptomatic/supportive treatment is the main treatment method,and some neonates tend to have a poor prognosis.

Clostridium difficile infection(CDI)is a major cause of hospital-acquired gastrointestinal infections in children.Current treatment for pediatric CDI primarily involves antibiotics;however,some children experience recurrence after antibiotic treatment,and those with initial recurrence remain at risk for further recurrences following subsequent antibiotic therapy.In such cases,careful consideration of treatment options is necessary.Fecal microbiota transplantation has been shown to be effective for recurrent CDI and has a high safety profile.This article reviews the latest research on the pathogenesis,risk factors,diagnosis,and treatment of pediatric CDI domestically and internationally,with a particular focus on fecal microbiota transplantation therapy.

Based on the interaction between supramolecule of traditional Chinese medicine and enterobacteria, the material basis of Rhei Radix et Rhizoma and Coptidis Rhizoma was explored. Scanning electron microscopy (SEM) and dynamic light scattering (DLS) were used to characterize the morphological differences of Rhubarb single decoction, Coptis single decoction and Rhubarb and Coptis co-decoction. An in vitro antibacterial model (E. coli, E. faecium and B. subtilis) was established to evaluate the damage effect of the combination of Rhei Radix et Rhizoma and Coptidis Rhizoma on enterobacteria. Ultra high performance liquid chromatography-tandem mass spectrometry (UPLC-MS/MS) was used to analyze the changes of chemical components of single decoctions and co-decoctions. The co-decoction of Rhei Radix et Rhizoma and Coptidis Rhizoma was turbid after decocting. The spherical particles of 300-400 nm were observed under SEM, and the co-decoction was more uniform and stable than that of single decoction. The interaction between supramolecules formed after the combination of Rhei Radix et Rhizoma and Coptidis Rhizoma and enterobacteria was significantly different from that of single decoction. In the process of interaction between supramolecules and enterobacteria, the spherical state was maintained, and the medicinal ingredients in Coptidis Rhizoma or Rhei Radix et Rhizoma were blocked, which could effectively alleviate the damage to enterobacteria. This study provided a reference for subsequent studies on the regulation of intestinal flora homeostasis by the combination of Rhei Radix et Rhizoma and Coptidis Rhizoma.

Tumor cells use glycolysis to provide material and energy under hypoxic conditions to meet the energy requirements for rapid growth and proliferation， namely the Warburg effect. Even under aerobic conditions， tumor cells mainly rely on glycolysis to provide energy. Therefore， glucose transporter protein 1（GLUT1）， which is involved in the process of glucose metabolism， plays an important role in tumorigenesis， development and drug resistance， and is considered to be one of the important targets in the treatment of malignant tumors. In recent years， research on tumor glucose metabolism has gradually become a hot spot. It has been shown that various factors are involved in the regulation of tumor energy metabolism， among which the role of GLUT1 is the most critical. In this paper， the authors reviewed the latest research progress of GLUT1-targeted traditional Chinese medicine（TCM） active ingredient nano-delivery system in tumor therapy， aiming to reveal the feasibility and effectiveness of this system in the delivery of chemotherapeutic drugs. The GLUT1-targeted TCM active ingredient nano-delivery system can overcome the bottleneck of the traditional targeting strategy as well as the high-permeability long retention（EPR） effect. In summary， the authors believe that the GLUT1-targeted TCM active ingredient nano-delivery system provides a new strategy for targeted treatment of tumors and has a broad application prospect in tumor prevention and treatment.

Licorice-gypsum (gancao-shigao, GC-SG) drug pair was used as the research object, using supramolecular chemistry to explore the scientific connotation of combining herbal medicine GC with insoluble mineral medicine SG in clinical application of traditional Chinese medicine. ① The Tyndall effect, microscopic morphology and particle size of the single and co-decocted of GC and SG were observed, the paste content and conductivity were determined, and the interaction between GC and SG was detected by isothermal titration calorimetry (ITC) and infrared absorption spectroscopy (IR). ② Calcium chloride (CaCl₂), a soluble calcium salt of equal gypsum quality, was used instead of SG with GC for co-decocting to explore the effect of calcium salt content on the water decocting, and the characteristics were combined with the Tyndall effect, microscopic morphology, paste content and conductivity. ITC and IR techniques were used to detect the interaction between the two, and the interaction between them was detected by ITC and IR. The zeta potential and ultraviolet-visible spectrophotometry (UV-vis) of GC-SG and GC-CaCl₂ co-decoction were compared, and the inorganic and organic components in the co-decoction were detected by inductively coupled plasma optical emission spectrometer (ICP-OES) and high performance liquid chromatography (HPLC). The results showed: ① Compared with the liquid phase of single decoction, GC-SG co-decoction had more obvious Tyndall effect, and showed uniform spherical nanoparticles under electron microscope. Physical characterization results such as paste content and conductivity showed that co-decoction promoted the dissolution of each other's components; ITC and IR results showed that there was strong interaction between GC and SG, which preliminatively indicated that GC and SG co-decoction promoted the formation of uniform and stable supramolecular system of traditional Chinese medicine. ② When soluble calcium salt was used to substitute insoluble SG with GC for co-decocting, a stronger but astigmatic light path appeared than single decocting solution, the zeta potential was reduced, and a large number of accumulated polymers were formed. The results of paste content and conductivity showed that the dissolution of the co-decocting component was reduced than the single decocting component. ITC, UV-vis and IR results showed that there was interaction between GC with Ca²⁺ and SG. The formation of polysink indicated that a large amount of soluble calcium salt would destroy the stability of supramolecular Chinese medicine. The results of ICP-OES and HPLC showed that the glycyrrhizic acid (GA) content of the former lower than the latter, which was related to the formation of a large number of polycondensates with the increase of Ca²⁺ concentration and the decrease of the dissolution of GA and other active ingredients. This study indicates that the compatibility of GC and SG can form a uniform and stable supramolecular system of traditional Chinese medicine. Calcium salt, the main component of SG, is taken as the starting point. Excessive soluble Ca²⁺ can promote the aggregation of active ingredients such as GA, so as to reveal the scientific connotation of the compatibility of GC and SG, an insoluble mineral medicine.

The traditional commodity specifications of Chinese medicinal materials are mainly divided into different grades based on macroscopic characteristics. As the basis for high quality and good price, there is still a lack of systematic evaluation on whether they are consistent with the current standards and whether they can reflect the internal quality of medicinal material. Panax notoginseng is a commonly used, large consumption of Chinese medicinal material. At present, it is divided into 8 grades in the market based on "Tou" (the number of crude drug /500 g), but it is not related to the standard of total saponins of Panax notoginseng (the sum of three saponins) in Chinese Pharmacopoeia. In this study, ultra-performance liquid chromatography-quadrupole-time of flight mass spectrometry (UPLC-Q-TOF-MS/MS) coupled with mass spectrometry molecular network were used for the rapid identification of saponins of Panax notoginseng with different "Tou" and a total of 64 saponins were identified. Seventeen saponins related to "Tou" were screened by orthogonal partial least squares discriminant analysis (OPLS-DA). The content of five saponins R₁, Rb₁, Rg₁, Rd, and Re in Panax notoginseng with different "Tou" was determined by high performance liquid chromatography (HPLC). The results of correlation analysis showed that Rd and R₁ with the largest VIP values among the differential saponins, which significantly negatively correlated with "Tou" (P < 0.05). Based on the determination results of 36 batches of samples, using Rd/Total Panax notoginseng saponins (TPNS) ratio ( > 0.08) as the index, Panax notoginseng can be divided into two grades: 20-60 "Tou" (superior) and 80-200 "Tou" (qualified). Based on the concept of "macroscopic characteristics and chemical profiling", this study integrates the non-targeted analysis and quantitative determination methods to provide a new strategy for quality evaluation of Panax notoginseng.