Aim To investigate the preventive and therapeutic effects of the mfat-1 gene therapy on exper-imental autoimmune encephalomyelitis in mice.Meth-ods mfat-1 gene therapy was used to render the host endogenous capability of producing ω-3 PUFAs,con-comitantly reduce the levels of ω-6 PUFAs,and change the proportion of ω-3/ω-6 PUFAs.Then,the levels of PUFAs in blood were analyzed by gas chromatography.The neurological deficits in mice were evaluated by neurological dysfunction score.HE staining and LFB staining of mouse spinal cord slices were used to ob-serve central nervous system inflammation infiltration and demyelinating lesions.Flow cytometry microsphere microarray technology was used to detect the content of cytokines in serum.Results The mfat-1 gene therapy could significantly raise the proportion of ω-3/ω-6 PU-FAs(P＜0.05),markedly delay the incubation period and peak period and reduce neurological dysfunction scores(P＜0.05),and improve inflammation and de-myelination of spinal cords(P＜0.05).It could also greatly increase the levels of IL-2,IFN-γ,IL-4 and IL-17 in serum(P＜0.05).Conclusion The pro-portion of ω-3/ω-6 PUFAs in blood circulation en-hanced by mfat-1 gene therapy can effectively prevent and treat experimental autoimmune encephalomyelitis in mice.

Coronary artery fistula(CAF)is defined as the direct connection between coronary artery and cardiac cavity,great vessel or other vascular structures.Most CAF are congenital and often present as persistent murmur in asymptomatic children.Although abnormal shunt of most coronary fistulas has little effect on hemodynamics,some larger fistulas can also lead to a series of complications,such as congestive heart failure.Therefore,closed CAF has been recommended to treat or prevent myocardial ischemia,congestive heart failure,endarteritis,arrhythmia,thromboembolism,aneurysm dilatation and rupture and other complications.CAF is a rare congenital heart disease,and bilateral CAF originating from two coronary arteries is even rarer.At present,there are rare systematic reports on bilateral CAF.Therefore,a case of huge bilateral CAF at the same site was reported,and systematically,this kind of case was summarized,so as to improve the clinical attention of diagnosis and treatment of this kind of CAF.Nowadays,some new interventional devices and technologies are emerging with the continuous progress of cardiovascular interventional technology,making the treatment more accurate,safe and effective.At the same time,there is also a clearer understanding of the indications and operating standards for interventional treatment,which is more operable and predictable during the treatment process.Therefore,interventional treatment of CAF may partially replace surgical treatment as the preferred treatment method.

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.

Since the outbreak of severe acute respiratory syndrome coronavirus 2(SARS-CoV-2)infection at the end of 2019,its global research has become a hot spot.The tendency of coronavirus infection mainly depends on the ability of spike protein(S protein)binding to receptors on the cell surface.S protein and its receptor binding domain(S-RBD)not only play a key role in the binding of virus to host cells and the entry of virus into cells,they can also bind to host cell surface receptors such as angiotensin-converting enzyme 2(ACE2),Toll-like receptor(TLRs),cluster of differentiation(CD)147,and neuropilin 1(NRP-1),activating different signaling pathways,thus promo-ting virus to invade host cells and trigger a series of pathogenic processes such as inflammation.Therefore,it is of great significance to study receptors involving in the interaction between S protein and host cells.This article re-views the receptors related to the interaction between S protein and host cells,so as to provide theoretical basis for the prevention and treatment of SARS-CoV-2 infection.

Aim To analyze the anti-A549 and HI299 lung ade-nocarcinoma activities via using examples of baicalin, astragalo-side, hesperidin and cisplatin based on real time cellular analysis (RTCA) technology, and to build a new strategy for EC50 e-valuation reflecting the time-dimensional characteristic. Methods Using RTCA Software Pro for data analysis and GraphPad Prism and Origin Pro plotting, the in vitro anti-A549 and H1299 lung adenocarcinoma activities of baicalin, astragaloside, hesperidin, and cisplatin were characterized using the endpoint method and time dimension, respectively. Results (X) There were significant differences in EC50 values of A549 and H1299 cells at 24 h and 48 h endpoint methods. (2) The correlation coefficient of the curve fitted with the four-parameter equation was > 0. 9, and the dynamic change of EC50 remained relatively stable (the linear fitting of EC50 at adjacent 4 points I slope 1^1) used to calculate the EC50 value within this time dimension. The EC50 of baicalin, astragaloside, hesperidin and cisplatin on A549 cells was 52. 97 ±1.75 плпо! • L~1(16~48 h) , 62.88 ± 2.91 ijunol • L"1 (32.25 -48 h) , 78.84 ±0.33 плпо1 • L"1 (21.5 -29.75 h), 13.57 ±1.54 плпо1 • L_1(27.5 -48 h), respectively; the EC50 of baicalin, astragaloside, hesperidin and cisplatin on H1299 cells was 43. 71 ± 1. 26 |лто1 • L_1 ( 19. 5 -48 h), 47.23 ±1. 19 |лто1 • L_1(14 -48 h) , 39.45 ±0.24 плпо1 • L"1 (12.75 -46.25 h), 25.97 ±4.76 плпо1 • L"1 (10. 25 -48 h) , respectively. The results showed that the time window for the anti-tumor effect of the test solution/drug was different. Conclusions Based on RTCA technology, it is more accurate and reasonable to select EC50 data that exhibit better fitting, stable changes, and time-dimensional characteristics for the evaluation of anti-tumor activity. In addition, this method of distinguishing different effective time of antitumor drugs can provide a reference for the timing of clinical combination drugs, and this approach will also provide a reference for further related studies.

Objective This study aims to investigate the impact of cultivation time on dendritic cells(DCs)and their derived exosomes′ expression of immune-related membrane proteins(CD80,MHC-Ⅰ,MHC-Ⅱ)and provides experimental evidence for future research.Methods Mouse bone marrow cells were induced to differentiate into DCs using GM-CSF and IL-4,followed by maturation stimulation withTNF-α.Exosomes were extracted using ultracentrifugation.Western blot and Amnis image flow cytometry were used to identify exosomes derived from mouse DCs.Amnis image flow cytometry was used to detect the expression of immune-related proteins CD80,CD11c,MHC-Ⅰ,and MHC-Ⅱ in mouse DCs and their exosomes.Results After 5 days of in vitro cultivation,more than 50%of dendritic cells expressed CD80,CD11c,MHC-Ⅰ,and MHC-Ⅱ,reaching the highest level on day 13.The positivity rates were as follows:CD80(97.29±0.63)%,CD11c(92.31±1.18)%,MHC-Ⅰ(97.91±0.49)%,and MHC-Ⅱ(97.91±0.49)%.The differences were statistically significant(P<0.001).The expression gradually decreased after day 13,but approximately 80%of DC cells still expressed MHC-Ⅰ and MHC-Ⅱ immune molecules on day 30.The expression levels of CD80,CD11c,and MHC-Ⅱ on the exosome membrane were highest on day 5 and then decreased overall with prolonged cultivation time,except for MHC-Ⅰ molecules.The differences were statistically significant(P<0.01).Conclusions In vitro-cultured mouse dendritic cells express high levels of immune-related membrane proteins and can be stably maintained for a long time under suitable culture conditions.The secreted exosomes also carry abundant immune-related membrane proteins,but no significant correlation was found between the immune-related proteins on the dendritic cell surface and the exosome membrane surface.

The covalent binding of drugs and their metabolites to proteins forms drug-protein adducts, which may cause adverse reactions in the body. The development of adductomics technology is helpful for the identification of covalent adducts between drugs and human plasma proteins. For many drugs, such as beta-lactam antibiotics, acyl glucuronides, covalent tyrosine kinases inhibitors, and reactive metabolites, human serum albumin (HSA) is a potential target and biomarker for the formation of drug-protein adducts. In this review, we will describe the relevant technical advances, describe the methods for the identification of covalent adducts of drugs and HSA, define the chemical reactions that form adducts, and preliminarily explore the role of drug-HSA adducts in adverse drug reactions and the potential effect on pharmacokinetics.

Objective To investigate the effect of microglia activation regulated by C-X3-C motif chemokine ligand 1(CX3CL1)-C-X3-C motif chemokine receptor 1(CX3CR1)pathway on memory function in hemorrhagic shock/resuscitation rats.Methods The experiment was divided into two parts.In the first part,the rats were randomly divided into sham group,model-0.5 hour group,model-1.5 hour group,model-3 hour group,10 rats in each group.There were differences in the time of hemorrhagic shock among each group.In the second part,rats were randomly divided into control group and CX3CL1 group,10 rats in each group.The rats in CX3CL1 group were treated with CX3CL1 protein factor(intraventricular injection),and the rats in control group were treated with saline.All rats were trained in Morris water maze experiments before model construction,and tests of Morris water maze experiments were carried out after 4 days of model construction.After completion,the whole brains were taken for HE staining and immunohistochemical staining.Cerebrospinal fluid was taken for detection of inflammatory cytokines,and hippocampus tissues were taken for Real-time PCR detection and Western blotting detection.Results Compared with the sham group,the escape latency of rats in model group increased,the number of platform crossings and the resident time in the third quadrant decreased.The neuronal state was impaired in HE staining in model group.In addition,compared with the sham group,the expression of ionized calcium binding adaptor molecule-1(Iba1)in the brain of the rats in model group increased,the contents of tumor necrosis factor-α(TNF-α)and interleukin(IL)-6 in the cerebrospinal fluid increased,and the M1-type microglia markers CD16,TNF-α,IL-1β and inducible nitric oxide synthase(iNOS)mRNA content increased.At the same time,compared with the sham group,the expressions of CX3CL1 and CX3CR1 in the brain of model group decreased,and the expressions of phosphorylated nuclear factor-κB(p-NF-κB)and nucleotide binding oligomerization domain(NOD)-like receptor protein 3(NLRP3)increased.However,compared with the control group,rats in CX3CL1 group had reduced escape latency,increased platform crossing times and quadrantⅢresident time,and recovered neuronal states.In addition,the expression of Iba1 in the brain of CX3CL1 group decreased,the contents of TNF-α and IL-6 in the cerebrospinal fluid decreased,the mRNA contents of M1-type microglia markers like CD16,TNF-α,IL-1β and iNOS decreased,and the mRNA contents of markers of M2-type microglia glial like CD206,transforming growth factor-β(TGF-β),arginase-1(Arg1),Chitinase 3-like protein 1(Ym 1)increased.Conclusion CX3CL1 can help inhibit the excessive activation of microglia,induce the polarization of microglia to M2 type,inhibit the polarization of M1 type,reduce the release of inflammatory cytokines,and alleviate the memory function damage induced by hemorrhagic shock/resuscitation.

Purpose::Vibrio vulnificus ( V. Vulnificus) infection is characterized by rapid onset, aggressive progression, and challenging treatment. Bacterial resistance poses a significant challenge for clinical anti-infection treatment and is thus the subject of research. Enhancing host infection tolerance represents a novel infection prevention strategy to improve patient survival. Our team initially identified cytochrome P4501A1 (CYP1A1) as an important target owing to its negative modulation of the body's infection tolerance. This study explored the superior effects of the CYP1A1 inhibitor bergamottin compared to antibiotic combination therapy on the survival of mice infected with multidrug-resistant V. Vulnificus and the protection of their vital organs. Methods::An increasing concentration gradient method was used to induce multidrug-resistant V. Vulnificus development. We established a lethal infection model in C57BL/6J male mice and evaluated the effect of bergamottin on mouse survival. A mild infection model was established in C57BL/6J male mice, and the serum levels of creatinine, urea nitrogen, aspartate aminotransferase, and alanine aminotransferase were determined using enzyme-linked immunosorbent assay to evaluate the effect of bergamottin on liver and kidney function. The morphological changes induced in the presence of bergamottin in mouse organs were evaluated by hematoxylin and eosin staining of liver and kidney tissues. The bacterial growth curve and organ load determination were used to evaluate whether bergamottin has a direct antibacterial effect on multidrug-resistant V. Vulnificus. Quantification of inflammatory factors in serum by enzyme-linked immunosorbent assay and the expression levels of inflammatory factors in liver and kidney tissues by real-time quantitative polymerase chain reaction were performed to evaluate the effect of bergamottin on inflammatory factor levels. Western blot analysis of IκBα, phosphorylated IκBα, p65, and phosphorylated p65 protein expression in liver and kidney tissues and in human hepatocellular carcinomas-2 and human kidney-2 cell lines was used to evaluate the effect of bergamottin on the nuclear factor kappa-B signaling pathway. One-way ANOVA and Kaplan-Meier analysis were used for statistical analysis. Results::In mice infected with multidrug-resistant V. Vulnificus, bergamottin prolonged survival ( p = 0.014), reduced the serum creatinine ( p = 0.002), urea nitrogen ( p = 0.030), aspartate aminotransferase ( p = 0.029), and alanine aminotransferase ( p = 0.003) levels, and protected the cellular morphology of liver and kidney tissues. Bergamottin inhibited interleukin (IL)-1β, IL-6, and tumor necrosis factor (TNF)-α expression in serum (IL-1β: p = 0.010, IL-6: p = 0.029, TNF-α: p = 0.025) and inhibited the protein expression of the inflammatory factors IL-1β, IL-6, TNF-α in liver (IL-1β: p = 0.010, IL-6: p = 0.011, TNF-α: p = 0.037) and kidney (IL-1β: p = 0.016, IL-6: p = 0.011, TNF-α: p = 0.008) tissues. Bergamottin did not affect the proliferation of multidrug-resistant V. Vulnificus or the bacterial load in the mouse peritoneal lavage fluid ( p = 0.225), liver ( p = 0.186), or kidney ( p = 0.637). Conclusion::Bergamottin enhances the tolerance of mice to multidrug-resistant V. Vulnificus infection. This study can serve as a reference and guide the development of novel clinical treatment strategies for V. Vulnificus.

Objective To investigate the role of magnesium ion(Mg2+)in cisplatin-induced acute kidney injury(Cis-AKI)in kidney organoids and HK-2 cells,as well as the potential mechanism.Methods Initially,we utilized human-derived induced pluripotent stem cells(iPSCs)to construct kidney organoids,and then built a Cis-AKI model based on kidney organoids.HE staining was used to observe the structure of kidney organoids,and immunofluorescence staining was used to observe the localization of markers and the expression of cleaved caspase-3.qRT-PCR was conducted to detect mRNA levels of tubular and glomerular markers,as well as inflammatory factors.Subsequently,the kidney organoids were randomly divided into control group,cisplatin group(Cis group),and Mg2+pretreatment group(Cis+Mg2+group).CCK-8 and ATP content assays were employed to evaluate the cell viability of renal tubular epithelial cells.TUNEL staining was performed to detect the apoptosis of renal tubular epithelial cells.Western blot was utilized to detect the expression of apoptosis-associated proteins(Bcl-2,Bax,cleaved caspase-3)and organic cation transporter 2(OCT2).Immunofluorescence was used to detect the localization and expression of OCT2.Results On the 10th day,the tubular structure in kidney organoids was visible,with abundant expression of renal markers.Treatment with 10 μmol/L cisplatin resulted in structural damage to kidney organoids,significantly increased expression of cleaved caspase-3 and mRNA levels of inflammatory factors,and significantly decreased ATP content.Compared with the Cis group,the Cis+Mg2+group showed increased ATP content in kidney organoids,reduced number of TUNEL-positive cells,significantly decreased expression of apoptosis-associated proteins,and significantly decreased expression of OCT2.However,there was no significant improvement in HK-2 cell viability,the number of TUNEL-positive cells,or apoptosis-associated proteins in the Cis+Mg2+group,and HK-2 cells did not express OCT2.Conclusion Kidney organoid is an ideal in vitro model to study the pathogenesis and treatment of Cis-AKI.Mg2+pretreatment can significantly reduce the damage of kidney organoids induced by cisplatin,and the mechanism may be related to the downregulation of OCT2.