Objective:To study the early predictors of refractory septic shock (RSS) in neonates.Methods:From July 2020 to December 2021, clinical data of neonates with septic shock admitted to the Neonatal Department of our hospital were retrospectively reviewed. According to the maximum septic shock score (SSS) during clinical course, the neonates were assigned into RSS group and non-RSS group. Perinatal data, laboratory results and hemodynamic parameters at diagnosis were compared between the two groups. Multiple logistic regression analysis was used to identify independent risk factors of RSS and septic shock-related death. Receiver operating characteristic (ROC) curve was constructed to evaluate the early predictors of poor prognosis.Results:A total of 130 neonates were enrolled, including 54 in RSS group and 76 in non-RSS group. Compared with the non-RSS group, the RSS group had significantly lower pH, base excess (BE), stroke volume index (SVI), cardiac output (CO) and cardiac index (CI).Meanwhile, the RSS group had significantly higher mean arterial pressure (MAP) to CI ratio (MAP/CI) and SSS [including bedside SSS (bSSS), computed SSS (cSSS) and modified version of cSSS (mcSSS)] (all P<0.05). Multiple logistic regression analysis showed that increased MAP/CI was an independent predictor of RSS. The cut-off value of MAP/CI was 11.6 [sensitivity 62%, specificity 87%, positive predictive value (PPV) 79% and negative predictive value (NPV) 77%], with an area under the curve (AUC) of 0.734. Increased mcSSS was an independent predictor of septic shock-related death. The cut-off value of mcSSS was 5.8 (sensitivity 83%, specificity 72%, PPV 21% and NPV 97%), with an AUC of 0.845. Conclusions:Increased MAP/CI (≥11.6) and mcSSS (≥5.8) may be early predictors of RSS and septic shock-related death in neonates.

Thromboembolic disease seriously affects people's health, and even endangers life. Nattokinase(NK) is an alkaline serine protease with strong thrombolytic activity and low toxicity. However, when NK passes through the stomach, it is degraded by gastric acid and pepsin, and subsequently loses its thrombolytic activity. The application of preparation technology can form a protective layer and improve the high bioavailability of NK. This article briefly introduced the pharmacological properties of NK, and discussed the characteristic of different dosage forms. The development of NK preparation was prospected in order to promote its research and application.

OBJECTIVE To investigate the protective effect and mechanism of quercetin on the cardiac and renal functions of rats with cardiorenal syndrome （CRS） based on the phosphoinositide 3-kinase （PI3K）/protein kinase B （Akt）/nuclear factor kappa- B （NF-κB） signaling pathway. METHODS CRS model of SD rats was induced by left anterior descending coronary artery ligation combined with acute renal ischemia-reperfusion. Model rats were randomly separated into model group， quercetin low-dose group （35 mg/kg）， quercetin high-dose group （70 mg/kg）， high-dose of quercetin+740Y-P group （70 mg/kg quercetin+3.5 mg/kg PI3K/Akt/ NF-κB signaling pathway activator 740Y-P）， with 12 rats in each group. Another 12 normal rats were selected as sham operation group. They were given relevant drugs， once a day， for 14 consecutive days. After administration， the cardiac function indexes [left ventricular ejection fraction （LVEF）， end-diastolic volume （EDV）， isovolumic relaxation time （IVRT）] and renal function indicators [blood urea nitrogen （BUN）， 24-hour urine protein， and serum creatinine （Scr）] were detected， and fibrosis in the cardiac and renal tissues was observed； the levels of inflammatory indexes [interleukin-1β （IL-1β） and tumor necrosis factor-α （TNF-α）] in the serum and cardiac and renal tissues as well as the expression of PI3K/Akt/NF-κB pathway-related proteins in the cardiac and renal tissues were detected. RESULTS Compared with sham operation group， the levels of BUN， 24-hour urine protein and Scr， collagen volume fraction of cardiac and renal tissues， the levels of IL-1β and TNF-α in serum and cardiac and renal tissues， and the phosphorylation of PI3K， Akt and NF-κB p65 protein in cardiac and renal tissues were increased significantly in model group （P＜0.05）； the levels of LVEF， IVRT and EDV were reduced significantly （P＜0.05）. Compared with the model group， the above indexes were reversed significantly in quercetin low-dose and high-dose groups （P＜0.05）， and the reversal effect was better in the high-dose group （P＜0.05）. 740Y-P restored the reverse effect of high-dose quercetin on the indexes （P＜0.05）. CONCLUSIONS Quercetin can alleviate cardiac and renal fibrosis and function injury， the mechanism of which may be 20232016） associated with inhibiting the activation of the PI3K/Akt/NF-κB signaling pathway.

ATG8-binding proteins play a key role in autophagy, selective autophagy or non-autophagy process by interacting between ATG8 and the ATG8-interacting motif (AIM) or the ubiquitin-interacting motif (UIM). There is great progress of ATG8-binding proteins in yeast and mammalian studies. However, the plant domain is still lagging behind. Therefore, the structure characteristics of plant ATG8 binding protein were firstly outlined. Unlike the single copy of ATG8 gene in yeast, many homologous genes have been identified in plant. The LIR/ AIM-docking site (LDS) of ATG8 protein contains W and L pockets and is responsible for binding to AIM. The ATG8 protein binds to UIM-containing proteins via UIM-docking site (UDS) instead of LDS. UDS is in the opposite position to LDS, so the ATG8 can bind both AIM and UIM proteins. Secondly, the structure and function of ATG8-binding proteins, especially the selective autophagy receptors, were systematically described. The protein NBR1 and Joka2, as proteaphagy receptors, guide ubiquitination protein aggregates to autophagosome for degradation by binding to AIM and ATG8 in Arabidopsis and tobacco, respectively. AtNBR1 also promotes plant immunity by binding the capsid protein of cauliflower mosaic virus and silencing suppressor HCpro of turnip mosaic virus, mediating pathogen autophagy. AtNBR1 still degrades chloroplast by microautophagy under photoinjure or chlorophagy during ibiotic stress. And the protein ORM mediates the degradation of plant immune receptor flagellin sensing 2 (FLS2) through AIM binding to ATG8. Interestingly, ATI1 and ATI2 participate in both chlorophagy and ERphagy. Otherwise, ER membrane protein AtSec62, soluble protein AtC53, and ubiquitin-fold modifier1-specific ligase 1 (UFL1) can be directly bound to ATG8 as ER autophagy receptors. As pexophagy receptor, AtPEX6 and AtPEX10 bind to ATG8 via AIM and participate in pexophagy. RPN10, as a 26S proteasome subunit, whose C-terminal UIM1 and UIM2 bind ubiquitin and ATG8, respectively, mediates the selective autophagy degradation of 26S proteasome inactivation when fully ubiquitinated. Plant-specific mitochondrial localization proteins FCS-like zinc finger (FLZ) and friendly (FMT) may also be mitophagy receptors. CLC2 binds to ATG8 via the AIM-LDS docking site and is recruited to autophagy degradation on the Golgi membrane. The tryptophan-rich sensory protein (TSPO) in Arabidopsis was involved in clearing free heme, porphyrin and plasma membrane intrinsic protein 2;7 (PIP2;7) through the combination of AIM and ATG8. The conformation of GSNOR1 changes during anoxia, exposing the interaction between AIM and ATG8, leading to selective degradation of GSNOR1. At last, the ATG8 binding proteins involved in autophagosome closure, transport and synthetic synthesis was summarized. For example, plant-specific FYVE domain protein required for endosomal sorting 1 (FREE1) is involved in the closure of autophagosomes during nutrient deficiency. Therefore, according to the recent research advances, the structure and function of plant ATG8-binding proteins were systematically summarized in this paper, in order to provide new ideas for the study of plant selective autophagy and autophagy.

The CRISPR/Cas system consists of clustered regularly interspaced short palindromic repeats (CRISPR) and CRISPR-associated genes (Cas). The system forms an adaptive immune system in archaea and bacteria. The inherent defense mechanism enables these microorganisms to protect themselves against the invasion of foreign genetic material. The system functions of immune response including three main stages: adaptation, expression/maturation, and interference, each stage needs specific Cas proteins encoded by Cas gene located near the CRISPR sequences, along with other auxiliary proteins. In 2015, Zhang et al. reportedCas12a (Cpf1) as a member of the Class II type V CRISPR/Cas12a system, which possesses endonuclease activity. This finding holds great promise for its application in the field of biotechnology. In 2018, Doudna’s team first applied the CRISPR/Cas12a system for detecting HPV nucleic acid. The system comprises the following essential components in vitro detection: Cas12a, the crRNA sequence complementary to the target DNA, the PAM sequence, and the ssDNA reporter. Cas12a possesses a typical RuvC domain, displaying a canonical bilobed architecture that consists of a recognition (REC) lobe and a nuclease (NUC) lobe. The REC lobe contains the REC1 and REC2 domains, and the NUC lobe includes RuvC, PAM-interacting (PI), Wedge (WED), and bridge helix (BH) domains. The mature crRNA for Cas12a has a length of 42-44 nt, consists of repeat sequence (19/20 nt) and spacer sequence (23-25 nt). The crRNA spacer sequence has been found to require a length of 18 nt to achieve complete cleavage activity in vitro. Additionally, mutation in the bases of crRNA can indeed affect the activity of Cas12a. The PAM sequence plays a critical role in the recognition and degradation of DNA by the CRISPR/Cas system, enabling the system to distinguish between self and non-self genomic materials. Cas12a can effectively target the spacer sequence downstream of a T-rich PAM sequence at the 5' end. LbCas12a and AsCas12a both recognize the PAM sequences of 5'-TTTN-3', while FnCas12a recognizes the PAM sequences of 5'-TTN-3'. All of these PAM sequences are located upstream on the non-template strand (NTS) at the 5' end. Cas12a (Cpf1), guided by the crRNA, binds to the target DNA by recognizing the PAM sequence. It exhibits the ability to induce arbitrary cleavage of ssDNA within the system while cleaving the target ssDNA or dsDNA. According to this feature, an array of nucleic acid detection methods has been developed for tumor detection and infection diagnostics, such as the DETECTR (RPA-CRISPR/Cas12a method) and HOLMES (PCR-CRISPR/Cas12a method) in 2018. Then, in 2019, Cas12aVDet (one-step detection method), where Cas12a protein was immobilized on the upper wall of the reaction tube. This not only prevented contamination from opening the tube but also reduced the detection reaction time. In 2021, the dWS-CRISPR (digital warm-start CRISPR) was developed as a one-pot detection method. It serves as an accurate approach for quantitatively detectingSARS-CoV-2 in clinical specimens. With the innovation of scientific technology, the high-sensitivity signal transduction technology has also been integrated with the CRISPR/Cas12a system, enabling direct detection of nucleic acids, and eliminating the need for nucleic acid amplification steps. Here, we elaborated the detection principles of CRISPR/Cas12a in in vitro detection. We discussed the different stages leading to the catalytic pathway of target DNA, and the practical applications of Cas12a in nucleic acid detection. These findings revealed a target interference mechanism that originates from the binding of Cas12a-guided RNA complex to complementary DNA sequences within PAM-dependent (dsDNA) regions. The crRNA-DNA binding activates Cas12a, enabling site-specific dsDNA cleavage and non-specific ssDNA trans-cleavage. The release of Cas12a ssDNase activity provides a novel approach to enhance the sensitivity and specificity of molecular diagnostic applications. Before these CRISPR/Cas12a-based nucleic acid detection methods can be introduced into clinical use, substantial work is still required to ensure the accuracy of diagnosis. Nevertheless, we believe that these innovative detection tools based on CRISPR/Cas will revolutionize future diagnostic technologies, particularly offering significant assistance in pathogen infection diagnosis for developing countries with relatively poor healthcare conditions and high prevalence of infectious diseases.

Objective:To investigate the association between congenital hypothyroidism (CH) and the adverse outcomes during hospitalization in very low birth weight infants (VLBWI).Methods:This prospective, multicenter observational cohort study was conducted based on the data from the Sino-northern Neonatal Network (SNN). Data of 5 818 VLBWI with birth weight <1 500 g and gestational age between 24-<37 weeks that were admitted to the 37 neonatal intensive care units from January 1 st, 2019 to December 31 st, 2022 were collected and analyzed. Thyroid function was first screened at 7 to 10 days after birth, followed by weekly tests within the first 4 weeks, and retested at 36 weeks of corrected gestational age or before discharge. The VLBWI were assigned to the CH group or non-CH group. Chi-square test, Fisher exact probability method, Wilcoxon rank sum test, univariate and multivariate Logistic regression were used to analyze the relationship between CH and poor prognosis during hospitalization in VLBWI. Results:A total of 5 818 eligible VLBWI were enrolled, with 2 982 (51.3%) males and the gestational age of 30 (29, 31) weeks. The incidence of CH was 5.5% (319 VLBWI). Among the CH group, only 121 VLBWI (37.9%) were diagnosed at the first screening. Univariate Logistic regression analysis showed that CH was associated with increased incidence of extrauterine growth retardation (EUGR) ( OR=1.31(1.04-1.64), P<0.05) and retinopathy of prematurity (ROP) of stage Ⅲ and above ( OR=1.74(1.11-2.75), P<0.05). However, multivariate Logistic regression analysis showed no significant correlation between CH and EUGR, moderate to severe bronchopulmonary dysplasia, grade Ⅲ to Ⅳ intraventricular hemorrhage, neonatal necrotizing enterocolitis in stage Ⅱ or above, and ROP in stage Ⅲ or above ( OR=1.04 (0.81-1.33), 0.79 (0.54-1.15), 1.15 (0.58-2.26), 1.43 (0.81-2.53), 1.12 (0.70-1.80), all P>0.05). Conclusion:There is no significant correlation between CH and in-hospital adverse outcomes, possibly due to timely diagnosis and active replacement therapy.

[Objective] To investigate the correlation between human leukocyte antigen (HLA) gene polymorphism and hepatitis B virus (HBV) infection. [Methods] Venous blood samples were collected from 501 healthy individuals undergoing physical examinations at Yan’an Hospital in Kunming, Yunnan Province. Enzyme linked immunosorbent assay (ELISA) was used to detect HBV halves. Based on the results of HBV half detection, the patients were divided into three groups: HBV carrier group, previous infection group, and healthy control group. The HLA-A antigen genotype was detected using polymerase chain reaction with sequence specific primers (PCR-SSP) genotyping technology, and the distribution frequency of HLA-A gene polymorphism was compared between HBV carrier group and healthy control group, as well as between previous infection group and healthy control group. SPSS17.0 software was used for data statistical analysis. [Results] In the healthy control group, the HLA-A2 positivity rate was 47.49%, and the allele frequency was 31.29%.The overall frequency of gene distribution in the healthy control group was consistent with the HLA-A allele table commonly and confirmed in China published by the Chinese Bone Marrow Bank. The HLA-A2 positivity rate and allele frequency in the HBV carrier group were 63.04% and 42.23%, respectively; The difference in HLA-A2 positivity rate and allele frequency among carriers was statistically significant (P<0.05). the HLA-A2 positivity rate and allele frequency in the HBV previous infection group were 56.14% and 35.97%, respectively, which did not significantly differ from those in the healthy control group (P>0.05). [Conclusion] HLA-A2 gene may be a susceptibility gene for chronic hepatitis B HBV carriers.

Objective:Operational efficiency and influencing factors of China's basic medical insurance system from 2020 to 2021 is conducted to provide reference for improving the operational efficiency and optimizing the input-output relationship.Methods:The super-efficiency SBM model based on unexpected output and the Malmquist index are used to measure the static and dynamic efficiency of resident medical insurance in 31 provinces in China,and Tobit regression analysis is employed to analyze the influencing factors.Results:The overall operational efficiency of resident medical insurance still needs improvement.The operational efficiency of resident medical insurance in the central and western regions is lower than that in the eastern region,and the gap is significant.Different levels and regions have differentiated main constraints on the operational efficiency of resident medical insurance.In terms of dynamic efficiency,the total factor productivity of resident medical insurance operation shows an increasing trend,mainly due to technological progress.In terms of influencing factors,the degree of aging,the level of medical expenses and the level of medical insurance supervision have a significant impact on the operational efficiency.Suggestions:Efforts should be made to bridge regional disparities,promote the equitable development of medical insurance,reasonably control the level of medical expenses,strengthen the supervision of medical insurance funds,and implement active aging policies.

Objective:To investigate the correlation between the expression of GLI1 and im-mune invasion and clinical prognosis in gastric cancer.To study the effect of GLI1 expression on drug resistance in gastric cancer.Methods:The expression difference of GLI1 in gastric cancer and normal tissues was analyzed by using TCGA database,and the effect of clinical features and GLI1 gene ex-pression level on prognosis of patients with gastric cancer was analyzed.The correlation between GLI1 gene expression and tumor immune cell infiltration in gastric cancer tissues was analyzed to explore its influence on drug resistance of chemotherapy drugs and targeted drugs.Clinical samples were collect-ed to analyze the difference of GLI1 expression in gastric cancer and paracancer tissues.Results:The expression of GLI1 in gastric cancer tissues was 1.7 times that in normal tissues,and the overall sur-vival and disease-free survival of patients with high expression are shorter than those with low ex-pression(P＜0.05).The interstitial score,immune score and abundance of immunoinfiltrating cells were higher in the high expression of GLI1 in gastric cancer tissues.High expression of GLI1 reduces drug sensitivity and is positively correlated with the expression of immune checkpoint markers PDCD1(P＜0.05).GLI1 expression was significantly increased in patients with subdifferentiated gastric cancer.Conclusions:GLI1 expression is associated with the prognosis and immune infiltration of patients with gastric cancer,and it may lead to poor prognosis of patients by regulating chemotherapy resis-tance,which may be a potential therapeutic target and molecular marker for gastric cancer.

The access evaluation of new medical technology is an important part of the preclinical application of medical technology and plays a vital role in ensuring the quality and safety of medical services.However,in the con-crete practice of access evaluation,there are still some problems such as imperfect access theoretical framework,imperfect evaluation index system.With the strategic support of health policies,laws,and regulations,the theory and method of HB-HTA are used for reference,core elements such as assessment subject,assessment object,and assessment content are comprehensively considered,the index system is designed from the dimensions of tech-nical characteristics,safety,effectiveness,economy and applicability,and the access evaluation framework of im-ported medical new technologies is constructed.To offer a theoretical framework and evidence-based basis for medi-cal facility medical technology access management.