Autism spectrum disorder （ASD）， also known as autism， is a series of neurodevelopmental disorders characterized by social disorders and repetitive stereotyped behaviors/narrow interests. Its pathogenesis is complex， and there is a lack of effective treatment drugs， with some cases having adverse outcomes. Recent studies have consistently revealed that individuals with autism spectrum disorder （ASD） commonly exhibit characteristics such as gut microbiota dysbiosis （abnormal Bacteroidetes/Firmicutes ratio）， impaired intestinal barrier function （elevated serum levels of zonulin and LPS）， and intestinal immune dysregulation （increased pro-inflammatory cytokines including IL-6 and TNF-α）， suggesting that gastrointestinal abnormalities may influence central nervous system development through neuroendocrine， immunoregulatory， and metabolic pathways. Consequently， growing scholarly attention has focused on dietary interventions as potential approaches to alleviate clinical symptoms in children with ASD. This review systematically summarizes the role of gut microbiota and their metabolite alterations in ASD pathogenesis， along with recent advancements in understanding the microbiota-gut-brain axis mechanisms. Additionally， it elaborates on the therapeutic effects and underlying biological basis of restrictive diet therapy， modified diet therapy， and nutritional supplementation therapy in promoting the health of children with ASD. This systematic review reveals that children with ASD exhibit significant gut microbiota dysbiosis （e.g.， increased Clostridium， decreased Faecalibacterium） and abnormal metabolite profiles （e.g.， altered short-chain fatty acid spectra， elevated 4EPS levels）. These alterations exacerbate neuroinflammation and immune dysregulation through the microbiota-gut-brain axis， thereby impacting nervous system development and function. Furthermore， interventions such as ketogenic diets， camel milk， and specific nutritional supplements can alleviate certain ASD symptoms by modulating gut microbiota， restoring intestinal barrier function， and improving metabolic pathways. Future investigations should aim to create multi-omics evaluation systems for pinpointing potential beneficiaries， devise individualized intervention strategies rooted in microbiome characteristics， and verify their therapeutic value and safety in large-scale randomized controlled trials. These efforts are crucial to transitioning ASD treatment from symptomatic control to address disease etiology， thereby paving the way for improving prognoses.

Transfusion and Anemia Expertise Initiative-Control/Avoidance of Bleeding developed a strategy for platelet and plasma infusion management in critically ill children based on systematic reviews and consensus meetings of international multidisciplinary experts. One good practice statement and six expert consensus statements were proposed for plasma and platelet transfusions in critically ill children following severe trauma, traumatic brain injury, and/or intracranial hemorrhage. This article introduces the specific methods and basis for the formation of recommendations in this part of the guide.

Eight compounds were isolated and purified from the ethyl acetate part of 70% acetone extract of Rehmannia glutinosa by various chromatographic techniques such as silica gel, MCI gel CHP-20, ODS, Toyopearl HW-40C, combined with TLC and semi-preparative HPLC. Their structures were elucidated by modern spectroscopy techniques (NMR, MS, UV, IR), and identified as neomartynoside A (1), osmanthuside B₆ (2), martynoside (3), isomartynoside (4), (E)-p-hydroxycinnamic acid (5), caffeic acid (6), ferulic acid (7), and methyl caffeate (8). Compound 1 is a new phenylethanol glycoside, which was identified as neomartynoside A. Compound 2 was isolated from Rehmannia glutinosa for the first time. In addition, compounds 2, 6 and 7 significantly increased relative glucose consumption, showing potential hypoglycemic activity.

ObjectiveTo investigate the effect of Qingrun prescription（QRP）-containing serum on improving insulin resistance in HepG2 cells and its potential mechanisms. MethodsAn insulin resistance model was established in HepG2 cells with 1×10^-6 mol·L^-1 insulin. Branched-chain α-keto acid dehydrogenase （BCKDH） gene silencing was achieved using siRNA， and the cells were divided into 8 groups： normal group， model group （1×10^-6 mol·L^-1 insulin）， metformin group （1 mmol·L^-1 metformin）， high-， medium-， and low-dose QRP groups （20%， 10%， and 5% QRP-containing serum， respectively）， QRP + siRNA-silenced BCKDH （si-BCKDH） group （10% QRP-containing serum + si-BCKDH）， and QRP + si-NC group （10% QRP-containing serum + si-NC）. Glucose levels in the supernatant were measured with a glucose assay kit， while glycogen content was assessed using a glycogen assay kit. Levels of branched-chain amino acids （BCAAs） and branched-chain keto acids （BCKAs） were determined using ultra-high performance liquid chromatography-tandem mass spectrometry （UHPLC-MS/MS）. mRNA transcription and protein expression levels of BCKDH， dishevelled， Egl-10， and pleckstrin （DEP） domain-containing mammalian target of rapamycin （mTOR）-interacting protein （DEPTOR）， mTOR， and ribosomal protein S6 kinase 1 （S6K1） were detected using real-time quantitative polymerase chain reaction （Real-time PCR） and Western blot. ResultsCompared to the normal group， the model group exhibited significantly decreased glucose consumption and glycogen content， increased levels of BCAAs and BCKAs， downregulated expression of BCKDH and DEPTOR， and upregulated mTOR and S6K1 expression （P<0.01）. In comparison to the model group， QRP treatment at all doses significantly enhanced glucose consumption and glycogen content while reducing BCAAs and BCKAs levels （P<0.01）. The high- and medium-dose QRP groups demonstrated significant upregulation of BCKDH mRNA transcription and protein expression， as well as DEPTOR mRNA transcription. Moreover， the DEPTOR protein expression level was significantly increased in high-， medium-， and low-dose QRP groups， while mTOR and S6K1 mRNA and protein expression levels were markedly downregulated （P<0.05， P<0.01）. Compared to the QRP + si-NC group， the QRP + si-BCKDH group exhibited increased BCAAs and BCKAs levels， significantly decreased BCKDH mRNA transcription and protein expression， downregulated DEPTOR mRNA and protein expression， and upregulated mTOR and S6K1 mRNA and protein expression （P<0.05， P<0.01）. ConclusionQRP may improve insulin resistance by reprogramming BCAAs metabolism. This effect involves upregulating BCKDH， reducing BCAAs and BCKAs levels， and suppressing the mTOR pathway activation.

Objective To establish a rapid,sensitive,and specific multiplex PCR detection method for the simultaneous detection of Cryptosporidium,Eimeria,and bovine parvovirus.Methods Specific primers targeting the SSU rRNA genes of Cryptosporidium and Eimeria,as well as the VP2 gene of bovine parvovirus were designed and the corresponding recombinant plasmid standards were constructed.To establish the multiplex PCR method,the reaction conditions were optimized using temperature gradient PCR and single-variable control methods.The sensitivity,specificity,reproducibility,and clinical application of the protocol were evaluated.Results The optimal annealing temperature was found to be 60.5℃,and the forward and reverse primer concentrations were determined to be 0.2 μmol/L for Eimeria,and 0.4 μmol/L for Cryptosporidium and bovine parvovirus.The assay demonstrated high sensitivity,with detection limits of 243,260,and 3 110 copies for the recombinant plasmid standards of Cryptosporidium,Eimeria,and bovine parvovirus,respectively.Specificity testing showed no cross-reactivity with ten common bovine pathogens,including Salmonella,bovine viral diarrhea virus,and bovine rotavirus.Consistent intra-and inter-batch results confirmed the strong reproducibility of the method.Clinical application to 81 diarrhea samples from various regions in the Ganzi Prefecture,Sichuan,revealed positivity rates of 18.52%(15/81)for Cryptosporidium,34.57%(28/81)for Eimeria,and 18.52%(15/81)forbovineparvovirus,withamixedinfectionrateof3.7%(3/81).Conclusions Themultiplex PCR method established in this study offers a reliable tool for differential diagnosis and epidemiological investigation of the three common diarrheal pathogens in yaks.

Objective To explore the traditional Chinese medicine(TCM)syndrome character-istics and medication patterns of cerebral small vessel disease(CSVD)using data mining techniques.Methods Clinical research literature on TCM treatment of CSVD,published from the establishment of the database to September 1,2024,was retrieved from the China National Knowledge Infrastructure(CNKI),Wanfang,and VIP databases.Analyses were conducted on syndromes,drug frequencies,properties,flavors,and meridian tropisms.Data mining was performed using R4.4.1 software to ex-plore associations,correlations,and clustering of TCM herbs,aiming to elucidate medication patterns in CSVD treatment.Results A total of 60 prescription formulas for treating CSVD were screened,involving 142 TCM herbs with a total usage frequency of 1,312 times.The top five most frequently used herbs were Chuanxiong,Danggui,Dilong,Huangqi,and Chishao.Herb properties were pre-dominantly warm and cold;flavors were mainly pungent,bitter,and sweet;and meridian tropisms were primarily to the liver,spleen,and heart meridians.Twenty-nine strong association rules were identified,and association analysis revealed core herbal combinations centered around Chuanxiong,Chishao,and Danggui.Clustering analysis yielded five herbal combinations.Conclusion CSVD is characterized by a deficiency in essence and excess in superficiality.Treatment should focus on tonify-ing deficiencies and eliminating excesses,combining both tonification and purgation methods.Medication patterns predominantly involve herbs for promoting blood circulation and removing blood stasis,often combined with herbs for nourishing the kidney and marrow,calming the liver and suppressing wind,and awakening the mind and opening the orifices.

Objective:To develop habitat radiomics models to predict early treatment responses to the hepatic arterial infusion chemotherapy (HAIC) combined with targeted therapy or immunotherapy in advanced hepatocellular carcinoma (HCC) patients, and to guide clinical diagnosis and treatment.Methods:From October 2021 to Decemeber 2023, at Army Characteristic Medical Center of PLA (Chongqing Daping Hospital) and the First Affiliated Hospital of Chongqing Medical University, 94 patients with advanced HCC who received HAIC combined with targeted therapy or immunotherapy were retrospectively enrolled. According to the treatment results, the patients were divided into response group and non-response group. Univariate and multivariate logistic regression were performed to analyze the clinical data of the patients. Based on contrast-enhanced CT images, tumor habitats were delineated and habitat features were extracted with k-means clustering, and the imaging features of arterial and venous phases were also extracted. The least absolute shrinkage and selection operator (LASSO) was used for dimensionality reduction. Feature selection was performed using LASSO to reduce dimensions, and then the selected features were further refined through stepwise logistic regression analysis.Binary logistic regression models were conducted to develop the habitat radiomics model, arterial phase radiomics model (APRM), venous phase radiomics model (VPRM), clinical data model, as well as the combination of radiomics model and clinical data model to predict early treatment (after 2 treatment cycles) response. Receiver operating characteristic curves (ROC) were plotted, and model performance was evaluated by the area under the curve (AUC), calibration curves, and decision curve. The models were validated through Bootstrap methods (1 000 times). DeLong test was used to compare AUC values.Results:The results of cluster analysis identified 3 characteristic habitats in HCC imaging: low-, medium-, and high-enhancement tumor habitats. The proportion of high-enhancement habitats was higher than that in the non-response group. A predictive model was established based on the proportions of these 3 habitats. Based on the proportion of low-, medium-, and high-enhancement habitats within the tumor, a habitat radiomics model was constructed. After LASSO selection and logistic regression analysis, 3 arterial phase and 3 venous phase radiomic features were selected to build the APRM and VPRM, respectively. Logistic regression analysis identified the following factors for the clinical data model: comorbidities ( OR=0.275, P=0.031), maximum tumor diameter ( OR=1.149, P=0.019), red blood cell count ( OR=0.463, P=0.022), alpha fetoprotein >400 μg/L ( OR=3.452, P=0.017), and tyrosine kinase inhibitor therapy ( OR=3.072, P=0.048). Among the single predictive model′s comparison, the AUC of habitat radiomics model was 0.860 (95% confidence interval(95% CI): 0.789 to 0.932), while those of the APRM、VPRM and clinical data model were 0.850 (95% CI: 0.773 to 0.926), 0.855 (95% CI: 0.782 to 0.928), and 0.774 (95% CI: 0.681 to 0.867), respectively, and there were no statistically significant among these models (all P>0.05). Among the combination models, the AUC of the habitat rediomic-clinical data combination model was 0.881 (95% CI: 0.814 to 0.947); the AUC of arterial phase rediomic-clinical data combination model was 0.897 (95% CI: 0.833 to 0.961); and the AUC of venous phase rediomic-clinical data combination model was 0.888 (95% CI: 0.826 to 0.951), but there were no statistically significant among the 3 models (all P>0.05). The calibration curve showed that the habitat rediomic-clinical data combination model had the most accurate predictive probability. Internal validation showed that the AUC of habitat rediomic-clinical data combination model was 0.848 (95% CI: 0.772 to 0.922), and the predictive performance was better than that of the clinical-data model (0.733 (95% CI: 0.670 to 0.863)). Conclusion:The habitat radiomics model based on enhanced CT can effectively predict early treatment responses to the HAIC combined with targeted therapy or immunotherapy in advanced HCC patients, which provides theoretical basis for individualized treatment in advanced HCC.

BACKGROUND:The cryopreservation technology enables tissues/cells to be stored for a long time in a low-temperature environment while maintaining the integrity of their activity and function,which is of great significance for the construction of cell therapy,tissue engineering and biological sample banks.Cryoprotective agents often contain dimethyl sulfoxide and serum.To avoid the toxic side effects of dimethyl sulfoxide,the complexity of serum components and immune responses,although some finished cryoprotective agents have been marketed,they are faced with many difficulties such as high cost and limited application.Therefore,there is an urgent need to develop a cryoprotective agent with clear components and the ability to solve the above problems.OBJECTIVE:To evaluate the effects of a novel cryoprotectant on cryopreservation efficiency of different tissue and cell sources.METHODS:By applying the novel cryoprotectant as an experimental group with the commercially available and widely used cryoprotectant(control group)to umbilical cord Wharton's jelly tissue,umbilical cord mesenchymal stem cells,umbilical cord blood/peripheral blood mononuclear cells,NK and CIK cells,comparative analyses were conducted in terms of cell morphology,number,viability,surface markers,differentiation potential,and cell-killing toxicity assay before cryopreservation and after resuscitation thawing.We confirmed the cryopreservation effect of the new cryoprotectant and its potential application value.RESULTS AND CONCLUSION:(1)The novel cryoprotectant facilitated the normal growth of cryopreserved Wharton's jelly tissue upon recovery,exhibiting mesenchymal stem cell morphology.No significant differences were observed between the experimental and control groups in terms of cell recovery rate,surface markers,and differentiation potential.(2)There was no significant difference in the number and viability of cells between the experimental group and the control group after cryopreservation of cord blood/peripheral blood mononuclear cells,and the cryo-resuscitated cell numbers and viability of derived NK cells/CIK cells did not show significant difference between the experimental and control groups.(3)For NK cells derived and differentiated from cord blood/peripheral blood mononuclear cells,there was no significant difference in the proportion of CD56+CD16+cell subpopulations between the experimental group and the control group.For CIK cells derived and differentiated from cord blood/peripheral blood mononuclear cells,there was no significant difference in the proportions of CD3+CD8+and CD3+CD56+cell subpopulations between the experimental group and the control group.(4)In terms of cytotoxicity testing,when the effective-target ratio of immune cells and melanoma cell line Mel624 was 20:1,whether it was NK cells/CIK cells derived from cord blood or peripheral blood mononuclear cells,there was no significant difference in the tumoricidal activity of cells between the experimental group and the control group.These findings suggest that the novel cryoprotectant can replace existing commercially available and widely used cryoprotectants,and is applicable to Wharton's jelly tissue,umbilical cord mesenchymal stem cells,umbilical cord blood/peripheral blood mononuclear cells,as well as NK and CIK cells,providing a solid technical foundation for the scaling,standardization,and commercialization of universal cryoprotectants.

A recombinant adenovirus(rAd)for expression of Mycobacterium tuberculosis(M.tb)c-di-AMP phosphodiesterase CnpB was constructed,and its induced humoral immune response was detected.The codon-optimized gene of M.tb CnpB was cloned into the adenoviral plasmid pcADV.The recombinant plasmid pcADV-CnpB was transfected into HEK293T cells,and expression was detected with Western blot.The recombinant plasmid pcADV-CnpB and the backbone plasmid were co-transfected into HEK293T cells to obtain the recombinant adenovirus rAd-CnpB.rAd-CnpB was amplified in HEK293T cells,and the target protein expression of rAd-CnpB was detected with Western blot and immunofluorescence.Mice were immunized with rAd-CnpB intranasally,and their sera and bronchoalveolar lavage fluid(BALF)were collected.ELISA was used to detect levels of antigen-specific antibodies.Restriction enzyme digestion and sequencing indicated that the recombinant plasmid pcADV-CnpB was successfully constructed and led to protein expression in eukaryotic cells.rAd-CnpB was packaged and produced in HEK293T cells.After amplification and purification,rAd-CnpB with a titer of 5.53×1010 PFU/mL was obtained.rAd-CnpB led to CnpB expression in HEK293T cells.Intranasal immunization with rAd-CnpB increased levels of IgG and secretory IgA in BALF and led to high levels of IgG in sera.rAd-CnpB,the recombinant adenovirus for expression of c-di-AMP phosphodiesterase CnpB was successfully constructed,and was found to induce antigen-specific humoral and mucosal immune responses through mucosal immunization.Thus,rAd-CnpB may be used in further research on new TB vaccine strategies.

This study assessed the role and mechanism of the recombinant Bacillus Calmette-Gue?rin vaccine(rBCG)with c-di-AMP adjuvant in regulating metabolism and immunity in epididymal white adipose(eWAT)in mice.Male C57BL/6 mice were intravenously immunized with BCG and rBCG,and their body weights were monitored.eWAT was isolated from the mice,and the stromal vascular fractions(SVFs)cell number was counted with a hemocytometer.Sections of mouse adipose tissue were prepared,and the size,number,and morphology of eWAT adipocytes and crown-like structure(CLS)formation were compared under a microscope after HE staining.The transcription levels of lipid metabolism-associated factors,cytokines and aging-associated genes in each group were determined with qRT-PCR.The body weights of mice gradually increased after immunization with BCG and rBCG.The proportions of eWAT increased,and the SVFs cell number decreased,in rBCG immunized mice.HE staining indicated that BCG immunization promoted hyperplasia,whereas rBCG immunization promoted hypertrophy of eWAT adipocytes;moreover,both BCG and rBCG immunization induced CLS formation in eWAT.The qRT-PCR results indicated that rBCG immunization inhibited the expression of genes associated with lipolysis and energy expenditure in eWAT.BCG immunization had little effect on cytokine transcription,whereas rBCG significantly induced the transcription of IFN-γ and IL-1Ra,and inhibited that of IL-15 and IL-2,but did not induce the expression of aging-associated genes.Thus,rBCG immunization induced eWAT adipocyte hypertrophy,which was associated with the inhibition of eWAT lipolysis and the regulation of cytokine expression.