The study explores the traditional Chinese medicine （TCM） diagnostic and treatment approach to coronary microvascular disease （CMVD） from the perspective of "disharmony of blood collaterals and dysfunction of qi transformation". It is proposed that the core pathogenesis of CMVD lies in these two mechanisms. From an integrative medicine perspective， different CMVD types are analyzed based on their specific pathogenesis. Through clinical practice， four targeted treatment methods， i.e. warming， unblocking， tonifying， and activating， are formulated. CMVD caused by atherosclerosis is primarily associated with myocardial ischemia， myocardial infarction， and coronary revascularization， with corresponding pathological mechanisms of latent pathogenic obstruction， toxic accumulation in the collaterals， and deficiency with collateral stasis. The disease progression exhibits characteristics of correlation， staging， and transformation. Accordingly， treatment principles include warming to assist qi transformation， unblocking obstruction and dispelling turbidity， activating to disperse toxic stasis and invigorate collaterals， and tonifying to eliminate stasis and nourish collaterals. For CMVD unrelated to atherosclerosis， attention should be paid to the underlying disease， analyzing the main syndromes of blood and collateral disharmony. An approach combining disease-syndrome differentiation with blood and collateral regulation is emphasized for precise treatment.

In protein engineering, while computational models are increasingly used to predict mutation effects, their evaluations primarily rely on high-throughput deep mutational scanning (DMS) experiments that use surrogate readouts, which may not adequately capture the complex biochemical properties of interest. Many proteins and their functions cannot be assessed through high-throughput methods due to technical limitations or the nature of the desired properties, and this is particularly true for the real industrial application scenario. Therefore, the desired testing datasets, will be small-size (∼10-100) experimental data for each protein, and involve as many proteins as possible and as many properties as possible, which is, however, lacking. Here, we present VenusMutHub, a comprehensive benchmark study using 905 small-scale experimental datasets curated from published literature and public databases, spanning 527 proteins across diverse functional properties including stability, activity, binding affinity, and selectivity. These datasets feature direct biochemical measurements rather than surrogate readouts, providing a more rigorous assessment of model performance in predicting mutations that affect specific molecular functions. We evaluate 23 computational models across various methodological paradigms, such as sequence-based, structure-informed and evolutionary approaches. This benchmark provides practical guidance for selecting appropriate prediction methods in protein engineering applications where accurate prediction of specific functional properties is crucial.

Objective To observe the impact of ultrasonic image quality on the consistency of artificial intelligence(Al)assisted diagnosis system and manual measurements of biological indicators of developmental dysplasia of hip(DDH).Methods Hip ultrasonic data of 75 DDH and 345 non-DDH children were retrospectively analyzed,and the quality of ultrasonic images were subjectively scored.An evaluation model of ultrasonic image quality was constructed based on 140 ultrasonic images acquired from 140 cases(group A,containing 25 DDH and 115 non-DDH)using entropy weighting method,the weight of anatomic structures and impact factors related to DDH were obtained.The comprehensive image quality scores of other ultrasonic images acquired from 280 cases(group B,including 50 DDH and 230 non-DDH)were calculated,and the images in group B were classified into grade A,B and C in descending order.The consistency of AI and manual measurements of DDH biological indicators in group B was assessed.Results The weight of each anatomic structure and impact factors of DDH obtained with the model were as follows:The lower edge of iliac branch＞ilium＞glenoid labrum＞bony margin＞femoral head＞motion artifacts.In group B,grade A was observed in 67(9 DDH and 58 non-DDH),grade B was found in 160(26 DDH and 134 non-DDH),while grade C was noticed in 53(15 DDH and 38 non DDH)images.Except for β,femoral head coverage(FHC)and femoral head length diameter,the consistencies between AI and manual measurements of other indicators of DDH were grade A＞B＞C.In group B,AI and manual measurements were more consistent in DDH than in non-DDH cases.Conclusion Ultrasonic image quality affected the consistency between AI and manual measurements of biological indicators of DDH.When image quality was not good enough,further attention should be paid to measurement of FHC and sizes of femoral head.

Single-cell or low-input multi-omics techniques have revolutionized the study of pre-implantation embryo development.However,the single-cell or low-input proteomic research in this field is rela-tively underdeveloped because of the higher threshold of the starting material for mammalian embryo samples and the lack of hypersensitive proteome technology.In this study,a comprehensive solution of ultrasensitive proteome technology(CS-UPT)was developed for single-cell or low-input mouse oocyte/embryo samples.The deep coverage and high-throughput routes significantly reduced the starting material and were selected by investigators based on their demands.Using the deep coverage route,we provided the first large-scale snapshot of the very early stage of mouse maternal-to-zygotic transition,including almost 5,500 protein groups from 20 mouse oocytes or zygotes for each sample.Moreover,significant protein regulatory networks centered on transcription factors and kinases between the MII oocyte and 1-cell embryo provided rich insights into minor zygotic genome activation.

Objective: To isolate a bacteriophage against pan-drug resistant Klebsiella pneumoniae in a burn patient, and to study its biological characteristics, genomic information, and effects on bacterial biofilm. Methods: (1) In 2018, pan-drug resistant Klebsiella pneumoniae UA168 (hereinafter referred to as the host bacteria) solution isolated from the blood of a burn patient in Ruijin Hospital Affiliated to Shanghai Jiao Tong University School of Medicine (hereinafter referred to as Ruijin Hospital) was used to isolate and purify the bacteriophage against pan-drug resistant Klebsiella pneumoniae from the sewage of Ruijin Hospital with sewage co-culture method, drip plate method, and double-agar plate method. The bacteriophage was named as phage KP168 and the plaque morphology was observed. (2) The phage KP168 solution was taken for cesium chloride density gradient centrifugation and dialysis, and then the morphology of phage KP168 was observed through transmission electron microscope after phosphotungstic acid negative staining. (3) The phage KP168 solution was taken to determine the lytic ability of the phage KP168 against 20 strains of pan-drug resistant Klebsiella pneumoniae isolated from the burned patients′ blood in Ruijin Hospital by the drip plate method, and then the lysis rate was calculated. (4) The phage KP168 solution at a initial titer of 9.3×10¹¹ plaque-forming unit (PFU)/mL (400 μL per tube) and the host bacteria solution at a concentration of 1×10⁹ colony-forming unit (CFU)/mL (4 mL per tube) were conventionally shaking cultured together for 4 hours at multiplicity of infection (MOI) of 10.000, 1.000, 0.100, 0.010, or 0.001, respectively (1 tube per MOI). The titer of phage KP168 was measured by the double-agar plate method (the measurement method was the same below) to select the optimal MOI. The experiment was repeated three times. (5) The host bacteria solution at a concentration of 1×10⁹ CFU/mL (4 mL per tube) and the phage KP168 solution at an adjusted titer of 5×10⁷ PFU/mL (400 μL per tube) were mixed at the MOI of 0.005. The plaques were counted 0 (immediately), 1, 2, 3, 4, 5, 15, and 30 minutes (1 tube at each time point) after mixing by the double-agar plate method (the counting method was the same below), and the percentage of adsorbed phages was calculated to screen for the optimal adsorption time. The experiment was repeated three times. (6) The host bacteria solution at a concentration of 1×10⁹ CFU/mL (300 μL per tube) and the phage KP168 solution at a titer of 5×10⁸ PFU/mL (60 μL per tube) were mixed at MOI of 0.005 and conventionally shaking cultured after standing for the optimal adsorption time. The phage KP168 titer was measured 0 (immediately), 10, 20, 30, 40, 50, 60, 70, 80, 90, and 100 minutes after culture, and a one-step growth curve was drawn. The experiment was repeated three times. (7) The phage KP168 solution at a titer of 2.5×10¹⁰ PFU/mL was left to stand for 1 hour at 37, 40, 50, 60, or 70 ℃ (3 tubes at each time point, 1 mL per tube) for counting the plaques, and then the thermal stability curve was drawn. SM buffer at a pH values of 5.0, 6.0, 7.0, 7.4, 8.0, 9.0, or 10.0 were added to the phage KP168 solution at a titer of 3.0×10¹⁰ PFU/mL, respectively. The mixed solution was left to stand for 1 hour at 37 ℃ (3 tubes of each pH, each tube containing 100 μL phage KP168 solution and 900 μL SM buffer), and then the plaques were counted, and an acid-base stability curve was drawn. (8) The phage KP168 solution was taken for DNA extraction and sequencing after dialysis as in experiment (2). The whole genome was annotated with Prokka to obtain the coding sequence of phage KP168. Nucleotide′s BLAST function was used to proceed nucleic acid sequence alignment for finding a known phage with the highest similarity to the phage KP168 nucleic acid sequence, and Blastx function was used to translate the coding sequence into protein for its function prediction. The comparison with Antibiotic Resistance Genes Database and Virulence Factors Database was proceeded. (9) In a 96-well plate, at a MOI of 1.000, 0.100, 0.010 or 0.001 (3 wells per MOI), 20 μL phage KP168 solution at a initial titer of 5.8×10¹⁰ PFU/mL was added to 200 μL host bacteria solution at a concentration of 1.5×10⁸ CFU/mL (the same concentration below) for co-cultivation for 48 hours. After 200 μL host bacteria solution was left to stand for 48 hours, 20 μL phage KP168 solution at a titer of 1×10^6, 1×10^7, 1×10^8, 1×10^9, or 1×10¹⁰ PFU/mL (3 wells per titer) was added respectively for action for 4 hours. In both experiments, 200 μL host bacteria solution added with 20 μL SM buffer (3 wells) acted as a negative control, and 220 μL LB culture medium (3 wells) acted as a blank control. Absorbance values were measured by a microplate reader, and inhibition/destruction rates of biofilm were calculated. The experiments were both repeated three times. Results: (1) The plaques of phage KP168 successfully isolated and purified were transparent and round, and its diameter was approximately 1.5 mm. (2) The phage KP168 has a regular polyhedron structure with a diameter of about 50 nm and without a tail. (3) The phage KP168 could lyse 13 of 20 strains of Klebsiella pneumoniae from burned patients, with a lysis rate of 65.0%. (4) When MOI was 1.000, the titer was the highest after co-culturing the phage KP168 with the host bacteria for 4 hours, which was the optimal MOI. (5) After the mixing of the phage KP168 with the host bacteria for 4 minutes, the percentage of the adsorbed phage reached the highest, which was the optimal adsorption time. (6) The one-step growth curve showed that during the lysis of the host bacteria by phage KP168, the incubation period was about 10 minutes, and the lysis period was about 40 minutes. (7) With the condition of 40 ℃ or pH 7.4, the number of plaques and the activity of phage KP168 reached the highest. (8) The genome of phage KP168 was a linear double-stranded DNA with a length of 40 114 bp. There were 48 possible coding sequences. It had the highest similarity to Klebsiella phage_vB_Kp1. The most similar known proteins corresponding to the translated proteins of coding sequences contained 23 hypothetical proteins and 25 proteins with known functions. No resistance genes or virulence factor genes were found. The GeneBank accession number was KT367885. (9) After 48 hours of co-cultivation of the phage KP168 and the host bacteria at each MOI, the inhibition rates of biofilm were similar, with an average of about 45%. After the phage KP168 with a titer of 1×10⁹ PFU/mL acted on the biofilm formed by the host bacteria for 4 h, the destruction rate of biofilm was the highest, reaching an average of 42%. Conclusions In this study, a bacteriophage against pan-drug resistant Klebsiella pneumoniae from a burn patient, phage KP168, is isolated from sewage, which belongs to the tailless phage. It has a wide host spectrum, short adsorption time, and short incubation period, with certain thermal and acid-base stability. Its genomic information is clear, and it does not contain resistance genes or virulence factor genes. It also has an inhibitory effect on the formation of bacterial biofilm and a destructive effect on the formed bacterial biofilm.

BRAF is a serine/threonine kinase that harbors activating mutations in ∼7% of human malignancies and ∼60% of melanomas. Despite initial clinical responses to BRAF inhibitors, patients frequently develop drug resistance. To identify candidate therapeutic targets for BRAF inhibitor resistant melanoma, we conduct CRISPR screens in melanoma cells harboring an activating BRAF mutation that had also acquired resistance to BRAF inhibitors. To investigate the mechanisms and pathways enabling resistance to BRAF inhibitors in melanomas, we integrate expression, ATAC-seq, and CRISPR screen data. We identify the JUN family transcription factors and the ETS family transcription factor ETV5 as key regulators of CDK6, which together enable resistance to BRAF inhibitors in melanoma cells. Our findings reveal genes contributing to resistance to a selective BRAF inhibitor PLX4720, providing new insights into gene regulation in BRAF inhibitor resistant melanoma cells.

Objective To investigate the reliability of using the pubic symphysis diastasis of 25 mm and anterior separation distance of sacroiliac joint to differentiate anteroposterior compression (APC) type Ⅰ and Ⅱ injuries as well as assess the injury severity.Methods A total of 11 (seven males and four females) fresh cadaver specimens with 22 hemipelvis were collected.The pelvic APC injury test models including fixed hemipelvis (restricted group) and unfixed hemipelvis (non-restricted group) were established,with 11 hemipelvis in each group according to the random number table method.Meanwhile the specimens were divided into male group (14 hemipelvis) and female group (eight hemipelvis),simulating APC type injury external rotation hemipelvis.The public symophysis interval and anterior interval of sacroiliac joint of the original pelvis,the pubic symphysis diastasis and anterior diastasis of sacroiliac joint after anterior tibiofibular ligament failure,as well as the affected pelvis ligament and sacral ligament injury were recorded and compared between the restricted and non-restricted groups,male and female groups.Results There were no significant differences in the public symphysis interval of the original pelvis and anterior interval of sacroiliac joint between the restricted group and the non-restricted group (P ＞ 0.05).The pubic symphysis interval of the original pelvis was [(5.13 ± 0.61) mm] in male group and (4.03 ± 0.84)mm] in female group (P ＜ 0.05).When the anterior tibiofibular ligament ruptured,the pubic symphysis diastasis distance was (23.36 ± 7.27) mm,ranging from 12 to 41 mm,and the diastasis distance of anterior sacroiliac joint was (9.82 ± 3.25)mm,ranging from 5 to 18 mm.In terms of the public symphysis interval,there were no significant differences between male and female groups,restricted and the non-restricted groups (P ＞ 0.05).In terms of anterior interval of sacroiliac joint,there was significant difference between male and female groups (P ＜ 0.05) but no significant difference between the restricted and non-restricted groups (P ＞ 0.05).In the restricted group,sacrotuberous ligament injuries were found in four patients,and sacrospinous ligament injuries in five,whhile there were no obvious sacrospinous ligament and sacrotuberous ligament injuries in non-restricted group.There were 10 specimens with the pubic symphysis diastasis ≥23.36 mm and 10 specimens with the diastasis distance of anterior sacroiliac joint ≥9.82 mm (46％),and there were 15 specimens with at least the pubic symphysis interval ≥ 23.36 mm or the anterior interval of sacroiliac joint ≥ 9.82 mm (68％).Conclusions The public symphysis interval ≥ 23.36 mm or anterior interval of sacroiliac joint ≥ 9.82 mm can distinguish anteroposterior compression Ⅰ from Ⅱ injuries,and the combination of the two criteria can be beneficial to assessment of pelvic injury severity.

Objectives To put forward the modified Letournel classification of acetabular fractures,and evaluate the guiding role of the modified Letournel classification in clinical work.Methods A retrospective study of 170 patients (178 sides) with acetabular fractures treated at Nanfang Hospital between January 2006 and August 2018 was performed.Among them 129 cases were males and 41 were females.The average age was 40.2±14.2 years (range,14-82 years).According to the Letournel classification,unclassifiable/atypical acetabular fractures were found out with plain Ⅹ-ray and plain CT scan and three-dimensional CT,and then their unclassifiable reasons were analyzed.Based on anatomical landmarks,clear boundaries of acetabular walls were defined on the 3D printed pelvic model.The structure of each column is a trihedron.The wall's fracture and column's fracture were distinguished using a theory that the wall's fracture involved two surfaces of column with interruption of continuity and column's fracture involved three.When column's fracture associated with wall's fracture,lowercase a,p and q were used representing comminuted areas of corresponding anterior and posterior wall,quadrilateral plate respectively.When column's fracture didn't associates with wall's fracture,it is marked with number 0.Finally,all fractures were classified according to the modified Letournel classification.The relationship between fracture type and surgical approach was analyzed.Results There was 51.7％ (92 sides) of acetabular fractures that couldn't be classified by the Letournel classification.Incomplete fracture lines (49 sides,53.3％),comminuted fractures (28 sides,30.4％),both of them (15 sides,16.3％) were their unclassifiable reasons.There were 8 types in the modified Letournel classification,including posterior wall fracture,posterior column fracture,anterior wall fracture,anterior column fracture,transverse fracture,T-shaped fracture,anterior column+posterior hemitransverse fracture,and both columns fracture.Posterior column+posterior wall fracture,transverse+posterior wall fracture had respectively become a form of posterior column fracture and of transverse fracture.According to the modified Letournel classification,the reclassification rate of 178 side fractures was 100％,which was significantly higher than 48.3％ (86/178) of the traditional Letournel classification,and the difference was statistically significant (x2=124.06,P＜ 0.001).100％ of posterior wall fracture and 80.00％ of posterior column fracture were treated by posterior approach.100％ of anterior column fracture and 73.68％ of both columns fracture were treated by anterior approach.Conclusion All acetabular fractures can be classified by the modified Letournel classification.The orientation of the comminuted walls reflects in part severity of the injury,better helps orthopaedic surgeons understand the morphology of acetabular fractures and select appropriate surgical approach.

Objective To establish a three-dimensional finite element model of pelvic anteroposterior compression (APC) for analysis of mechanisms for related liganentous damages.Methods A finite element model and a laboratory mechanical model of APC were established using the same pelvic specimens.In a finite element model of normal pelvic bones and ligaments,after the right pelvis was fixated the pubic symphysis (PS) was sectioned.Next,a manual external mobile force was gradually applied to the left hemipelvis to make the PS diastasis 10,20,30,40,60,80 and 100 mm apart.The mechanical experiment revealed the anterior sacroiliac ligament (ASIL) was ruptured when the PS diastasis reached 28 mm.After the strain value of ASIL was calculated through the finite element model,it was applied to the other pelvic ligaments.The displacement in front of the sacroiliac joint (SIJ),stress,strain and extent of injury and disruption of sacrotuberous/sacrospinous ligaments (STL/SSL) with a corresponding PS diastasis were observed and recorded.Results ASIL failed at the point when the PS diastasis was 28 mm and the displacement in front of SIJ was 7.41 ± 1.14 mm.The strain and maximum principal stress of ASIL calculated in the finite element model were 259.5％ and 543.24 MPa respectively.The maximum principal stress value of SSL was 35.00 MPa at the point of failure when the PS diastasis and the displacement in front of SIJ were 51 mm and 15.23 ±2.88 mm,respectively.When the PS diastasis and the displacement in front of SIJ were 100 mm and 7.5 mm respectively,the maximum principal stress value of STL was 16.17 MPa but the strained ligament was not ruptured.When the pelvis was rotated externally step by step,the ASIL failure was followed by the rupture of SSL but not necessarily by the STL failure.Conclusion As the finite element pelvic bone-ligament model established in this study can effectively simulate the mechanisms for APC injury,it can be used to evaluate different extents of pelvic ligamentous injury,providing a basis for the biomechanical study of pelvic bones and ligaments.

Objective To investigate the biomechanical stability of 5 kinds of internal fixation for acetabular T-shaped fractures using finite element analysis.Methods After a three-dimensional model of normal pelvis was reconstructed using Mimics software,the model of acetabular T-shaped fracture was divided by Solidworks software.Models of acetabular T-shaped fracture fixated by 5 methods were reconstructed:(A) a reconstruction plate for anterior column + lag screws for posterior column,(B) lag screws for anterior column + a reconstruction plate for posterior column,(C) a reconstruction plate for anterior column + a medial ilioischial plate for posterior column (combination plating),(D) retrograde lag screws for 2 columns,and (E) a reconstruction plate for anterior column + and a posterior ilioischial plate for posterior column.Valuation was conducted with software Ansys to simulate 600 N loading on the terminal plate of S1 vertebral body at standing and sitting positions.The biomechanical stabilities were compared between the 5 models of internal fixation.Results At standing and sitting positions,the displacements of anterior column in modelA (0.522±0.121 mmand0.762±0.181 mm),modelC (0.512±0.207mmand0.730±0.181mm) and model E (0.513 ± 0.184 mm and 0.733 ± 0.166 mm) were significantly smaller than those in model B (0.622±0.224 mmand 1.328±0.537 mm) and in modelD (0.655±0.174mm and 1.591± 0.270 mm) (P ＜ 0.05).However,there were no statistically significant differences in anterior column displacements between models A,C and E or between models B and D (P ＞ 0.05).At standing and sitting positions,the displacements of posterior column in model B (0.631 ± 0.151 mm and 0.572 ± 0.693 mm),in model C (0.621 ± 0.195 mm and 0.538 ± 0.075 mm) and in model E (0.625 ± 0.192 mm and 0.544 ± 0.063 mm) were significantly smaller than those in model A (0.742 ±0.140 mm and 0.715 ±0.072 mm)and in modelD (0.754±0.247 mm and 0.717 ± 0.071 mm) (P ＜0.05).However,there were no statistically significant differences in posterior column displacements between models B,C and E or between models A and D (P ＞ 0.05).Conclusions In the treatment of acetabular T-shaped fractures,reconstruction plating has better stability than screwing.Reconstructive plating shows the best stability for anterior and posterior columns,followed by combination plating which is the best choice for reducing surgical injury.