Acute-on-chronic liver failure （ACLF） is a syndrome of acute liver function decompensation in patients with chronic liver disease and is characterized by a high short-term mortality rate. Liver transplantation is an effective radical treatment method for ACLF and can significantly improve the survival rate of patients. However， in the context of a shortage of donor organs globally， how to achieve a balance between saving lives and avoiding futile transplantation while optimizing organ utilization is a critical challenge in clinical decision-making. This article systematically reviews the evolution of the evaluation and decision-making system for liver transplantation in ACLF patients and points out s that the core of this system has shifted from relying solely on the MELD score to a model based on multidimensional prognostic assessment， dynamic evaluation， and multidisciplinary collaboration. In the future， the integration of Eastern and Western criteria， the application of artificial intelligence for precise prediction， and the establishment of a more equitable priority and allocation system will be the key directions for development in this field.

Carotid artery perivascular adipose tissue (PVAT) can influence plaque formation and progression. Recently, carotid artery PVAT density has emerged as a novel imaging biomarker being capable of reflecting local metabolic and inflammatory states of adipose tissue. It is closely associated with vulnerable plaque characteristics, such as intraplaque hemorrhage, thinning or rupture of the fibrous cap, lipid-rich necrotic core, and calcification. Therefore, carotid artery PVAT density holds promise as a key parameter for early identification of vulnerable carotid plaques and stroke risk prediction. This article reviews the definition and pathophysiological mechanism of PVAT and application of imaging techniques in PVAT, as well as the association between carotid artery PVAT density and vulnerable characteristics of plaques, with the aim of providing references for early identification of asymptomatic high-risk plaques and individualized prevention strategies of ischemic stroke.

Objective:To evaluate the left atrial structure and function in patients with non-obstructive hypertrophic cardiomyopathy（NOHCM）and atrial fibrillation（AF）using four-dimensional automatic left atrial quantification（4D Auto LAQ）technology，and to explore the correlation between left atrial remodeling and AF.Methods:Ninety patients with NOHCM treated in the First Affiliated Hospital of Zhengzhou University from May 2023 to September 2024 were prospectively selected and divided into AF group（ n=27）and non-AF group（ n=63）according to whether they had AF. Clinical data of all enrolled patients were collected，and two-dimensional，color Doppler，and tissue Doppler ultrasound parameters were measured. 4D Auto LAQ technology was used to obtain left atrial volume and strain parameters. Inter-group comparison，univariate and multivariate Logistic regression analysis，and repeatability test were used to analyze the data. Results:Compared with non-AF group，left atrial maximum volume index（LAVImax），left atrial minimum volume index（LAVImin）and left atrial pre-systolic volume index（LAVIpreA）were increased in the AF group，but left atrial total ejection fraction（LAEF），left atrial passive ejection fraction（LApEF），left atrial active ejection fraction（LAaEF）and the absolute values of left atrial reservoir longitudinal and circumferential strains（LASr，LASr-c），left atrial conduit longitudinal and circumferential strains（LAScd，LAScd-c）and left atrial contraction longitudinal and circumferential strains（LASct，LASct-c）were decreased（all P<0.05）. Multivariate Logistic regression analysis showed that LAVImin was independently associated with AF（ OR=0.837， P<0.001）. Conclusions:4D Auto LAQ can quantitatively evaluate the left atrial structure and function in patients with NOHCM and AF，and LAVImin is independently associated with the presence of AF.

ObjectiveTo investigate the dynamic trajectories of prognostic scores and key clinical indicators in hepatitis B virus-related acute-on-chronic liver failure （HBV-ACLF） patients without liver cirrhosis， to clarify their association with outcomes， and to provide new evidence for individualized prognostic assessment. MethodsA prospective study was conducted for the data of 154 non-cirrhotic HBV-ACLF patients who attended Beijing YouAn Hospital of Capital Medical University from January 2016 to December 2023， including prognostic scores and key biochemical indicators on Days 3， 7， 14， 21， and 28 of the disease course. According to the outcome of patients at 1 year， they were divided into death/liver transplantation group with 43 patients， liver cirrhosis group with 23 patients， and non-liver cirrhosis group with 88 patients， and the trajectory heterogeneity of different outcome subgroups was analyzed. A one-way analysis of variance was used for comparison of normally distributed continuous data among the three groups； the Kruskal-Wallis H test was used for comparison of non-normally distributed continuous data among the three groups； the Wilcoxon test was used between two groups. the chi-square test was used for comparison of categorical data between groups. The mean and its 95% confidence interval （CI） were calculated for each indicator at difference time points； the linear interpolation method was used to connect the means at adjacent time points and construct the group-specific longitudinal trend curve； the 95%CI was visualized using the semi-transparent ribbon area， with the transparency parameter （α=0.2） optimized to enhance the visual discrimination of overlapping intervals across multiple groups. A linear mixed-effects model was used to compare the longitudinal changing trend of each indicator between the patients with different outcomes； likelihood ratio was used to evaluate the significance of the interaction effect between time and group， and in case of the significant interaction effect， the slope based on the estimated marginal mean was used for comparison between two groups. ResultsThere were significant differences between the three groups in the incidence rates of ascites and grade Ⅲ — Ⅳ hepatic encephalopathy， MELD score， MELD-Na score， CLIF-C ACLF score， COSSH-ACLF Ⅱ score， total bilirubin （TBil）， international normalized ratio （INR）， alpha-fetoprotein， blood sodium， alanine aminotransferase， and procalcitonin at the baseline（all P0.05）. The analysis of dynamic trajectories showed that the death/liver transplantation group had high levels of prognostic scores and the biochemical parameters of TBil and INR （TBil400 μmol/L， INR2.5）， as well as a low level of platelet count （PLT） （100×10⁹/L）. The non-liver cirrhosis group had rapid improvements in indicators， with TBil200 μmol/L， INR1.5， and PLT100×10⁹/L by day 28， while the liver cirrhosis group showed a trend of recovery， with TBil200 μmol/L， INR2.0， and PLT 100×10⁹/L on day 28， with significant global heterogeneity in the temporal trends of the above indicators across the three groups （all P0.01）. ConclusionDynamic monitoring of prognostic scores and key clinical indicators can effectively stratify the 1-year outcomes of non-cirrhotic patients with HBV-ACLF. Patients with poor prognosis were typically characterized by INR 2.5 and TBil 400 μmol/L. Among those who survived beyond 1 year， individuals who subsequently progressed to cirrhosis were frequently identified by the presence of INR 1.5， TBil 200 μmol/L， and PLT 100×10⁹/L at day 28.

Objective To assess the nutritional status in elderly patients with chronic renal insufficiency (CRI) and reveal the key factors affecting the nutritional status. Methods A total of 310 elderly patients with CRI who received hospitalization treatment and outpatient follow-up in the hospital from January 2021 to June 2024 were selected as the investigation subjects. The nutritional status of patients was evaluated by mini-nutritional assessment (MNA) questionnaire, and the nutritional status and dietary structure of patients were comprehensively evaluated by anthropometric indicators [height, weight, body mass index (BMI), upper arm circumference, calf circumference], biochemical indicators [serum albumin (ALB), prealbumin (PA), hemoglobin (Hb), transferrin (TF)] and 24-hour dietary review method. According to the investigation results of nutritional status, the patients were divided into good nutrition group (MNA score≥24 points), nutritional risk group (MNA score of 17-23.5 points) and malnutrition group (MNA score<17 points). Univariate analysis was adopted to screen the potential influencing factors of elderly CRI. Multivariate logistic regression model was applied to analyze the influencing factors of malnutrition in elderly CRI patients. Results Among the 325 questionnaires were distributed, but only 310 valid questionnaires were recovered, with an effective recovery rate of 95.38%. Investigation results revealed that among the 310 patients, 29.35% (91 cases) had good nutritional status, and 42.26% (131 cases) had nutritional risk, and 28.39% (88 cases) had malnutrition. Univariate analysis indicated that there were statistical differences in BMI, CRI staging, serum ALB, PA, Hb, TF, protein intake and total calorie intake among the good nutrition group, the nutritional risk group and the malnutrition group (P<0.05). Multivariate logistic regression analysis suggested that low BMI (OR=0.903, 95%CI: 0.867-0.941), high CRI stage (OR=1.091, 95%CI: 1.053-1.130), low serum ALB (OR=0.907, 95%CI: 0.867-0.948), PA (OR=0.918, 95%CI: 0.888-0.949), Hb (OR=0.944, 95%CI: 0.909-0.997), TF (OR=0.912, 95%CI: 0.874-0.952), insufficient protein intake (OR=0.924, 95%CI: 0.882-0.969) and insufficient total calorie intake (OR=0.938, 95%CI: 0.909-0.968) were influencing factors for malnutrition in elderly patients with CRI (all P<0.05). Drawing ROC curve of malnutrition in elderly patients with CRI according to the prediction probability of logistic regression model found that the AUC, sensitivity, specificity, 95%CI and Youden index were 0.976, 93.18%, 92.34%, 0.953-0.990 (P<0.05) and 0.855. Conclusion The incidence rate of malnutrition is high in elderly patients with CRI, and is mainly affected by factors such as low BMI, high CRI stage, low serum ALB, PA, Hb and TF levels and insufficient protein and total calorie intakes. In addition, logistic regression model has high predictive value and can provide a reference for early clinical identification of high-risk population with malnutrition among elderly patients with CRI.

Selection markers are essential tools in gene editing, the utility of such systems is inherently constrained by species-specific limitations, governed by divergent host genetic backgrounds and metabolic compatibility. To address this limitation, we leveraged the CRISPR/Cas9 system to develop a universal counter-selection tool. We designed and introduced an sgRNA expression cassettes as counter-selection markers, which directs the Cas9 protein to target and cleave genomic DNA, allowing for the selection of the strains where the sgRNA expression cassette has been replaced. Optimized to target multiple copy sites with sgRNA, this system significantly enhances cell lethality, boosting counter-selection efficiency to over 85.00%. This counter-selection tool is not limited to single strains and is suitable for various scenarios, including multi-copy plasmid assembly and plasmid editing, demonstrating broad application potential.
CRISPR-Cas Systems/genetics* ; Gene Editing/methods* ; RNA, Guide, CRISPR-Cas Systems/genetics* ; Plasmids/genetics*

Objective : To investigate the expression of Keratin 14(KRT14) in Basal-like Breast Cancer(BLBC) and its biological functions and mechanisms. Methods : The expression levels of KRT14 mRNA in BLBC and para-cancer breast tissues were analyzed using The Cancer Genome Atlas(TCGA) database. qPCR, Western blot(WB), and immunohistochemistry were employed to detect KRT14 expression in BLBC and adjacent normal tissues, and its correlation with clinicopathological features was analyzed. KRT14 overexpression and knockdown were performed in breast cancer cells, and cell scratch and transwell assays were performed to evaluate changes in migration and invasion abilities. To investigate the expression of proteins related to the Wnt/β-catenin signaling pathway, including catenin Beta 1(β-catenin), wingless-type MMTV integration site family, member 1(Wnt1), matrix metallopeptidase 7(MMP7), and cellular myelocytomatosis viral oncogene homolog(c-Myc), as well as the cellular localization of β-catenin, WB and immunofluorescence(IF) techniques were employed. Additionally, a Wnt/β-catenin signaling pathway inhibitor was used to verify the mechanism of action of KRT14. Results : The expression of KRT14 was significantly higher in BLBC tissues compared to normal tissues(P<0.05), and was associated with higher T stage and histological grade(P<0.05). The overexpression of KRT14 significantly enhanced the migration and invasion abilities of breast cancer cells, while the knockdown of KRT14 significantly reduced those abilities(P<0.01). The overexpression of KRT14 can increase the expression levels of Wnt/β-catenin pathway-related proteins β-catenin, Wnt1, MMP7, and c-Myc, thereby activating the Wnt/β-catenin pathway. Moreover, the inhibition of this pathway can eliminate the effects of KRT14 on cell migration and invasion. Conclusion The high expression of KRT14 in BLBC may promote the migration and invasion of breast cancer cells through the Wnt/β-catenin signaling pathway.

Objective : To investigate the expression of(heat shock protein a member 8,HSPA8) in breast cancer and its effect on tumor biological behaviors. Methods: Bioinformatics analysis and immunohistochemistry assays were used to detect the expression of HSPA8 in breast cancer and adjacent non-tumor breast tissues,and the relationship between its expression and clinicopathological characteristics of breast cancer patients was analyzed.Correlation between HSPA8 expression and prognosis of breast cancer patients was analyzed by Kaplan-Meier Plotter database.HSPA8 knockdown and over expression breast cancer stabilized cells were constructed,respectively.CCK-8,clone formation,Transwell,cell scratch,Western blot and immunofluorescence assay were used to detect the effects of HSPA8 on the proliferation,invasion and migration of breast cancer cells,and its effect on epithelial-mesenchymal transition(EMT). Results : Bioinformatics analysis and immunohistochemistry assay revealed that the expression of HSPA8 in breast cancer tissues was higher than that in adjacent non-tumour tissues(P<0.05),and its expression level of the protein was significantly and positively correlated with the tumor size,histological grade,lymph node metastasis and Ki-67 proliferation index(P<0.05).The Kaplan-Meier survival curve showed that high expression of HSPA8 was significantly associated with poor prognosis in breast cancer patients(P<0.05).CCK-8,clone formation,transwell,cell scratch,Western blot and immunofluorescence assay showed that knockdown of HSPA8 expression could significantly inhibit the proliferation,invasion,migration function and EMT of breast cancer cells(P<0.05),while overexpression of HSPA8 could significantly promote the proliferation,invasion,migration function and EMT of breast cancer cells(P<0.05). Conclusion HSPA8 is highly expressed in breast cancer tissues,which is closely related to disease progression and the malignant phenotype of breast cancer,suggesting that HSPA8 may be a potential biological target for breast cancer treatment.

Objective:To evaluate the left atrial structure and function in patients with non-obstructive hypertrophic cardiomyopathy（NOHCM）and atrial fibrillation（AF）using four-dimensional automatic left atrial quantification（4D Auto LAQ）technology，and to explore the correlation between left atrial remodeling and AF.Methods:Ninety patients with NOHCM treated in the First Affiliated Hospital of Zhengzhou University from May 2023 to September 2024 were prospectively selected and divided into AF group（ n=27）and non-AF group（ n=63）according to whether they had AF. Clinical data of all enrolled patients were collected，and two-dimensional，color Doppler，and tissue Doppler ultrasound parameters were measured. 4D Auto LAQ technology was used to obtain left atrial volume and strain parameters. Inter-group comparison，univariate and multivariate Logistic regression analysis，and repeatability test were used to analyze the data. Results:Compared with non-AF group，left atrial maximum volume index（LAVImax），left atrial minimum volume index（LAVImin）and left atrial pre-systolic volume index（LAVIpreA）were increased in the AF group，but left atrial total ejection fraction（LAEF），left atrial passive ejection fraction（LApEF），left atrial active ejection fraction（LAaEF）and the absolute values of left atrial reservoir longitudinal and circumferential strains（LASr，LASr-c），left atrial conduit longitudinal and circumferential strains（LAScd，LAScd-c）and left atrial contraction longitudinal and circumferential strains（LASct，LASct-c）were decreased（all P<0.05）. Multivariate Logistic regression analysis showed that LAVImin was independently associated with AF（ OR=0.837， P<0.001）. Conclusions:4D Auto LAQ can quantitatively evaluate the left atrial structure and function in patients with NOHCM and AF，and LAVImin is independently associated with the presence of AF.

Carotid artery perivascular adipose tissue (PVAT) can influence plaque formation and progression. Recently, carotid artery PVAT density has emerged as a novel imaging biomarker being capable of reflecting local metabolic and inflammatory states of adipose tissue. It is closely associated with vulnerable plaque characteristics, such as intraplaque hemorrhage, thinning or rupture of the fibrous cap, lipid-rich necrotic core, and calcification. Therefore, carotid artery PVAT density holds promise as a key parameter for early identification of vulnerable carotid plaques and stroke risk prediction. This article reviews the definition and pathophysiological mechanism of PVAT and application of imaging techniques in PVAT, as well as the association between carotid artery PVAT density and vulnerable characteristics of plaques, with the aim of providing references for early identification of asymptomatic high-risk plaques and individualized prevention strategies of ischemic stroke.