Objective To explore the clinical characteristics of patients with diffuse large B-cell lymphoma (DLBCL) accompanied with KMT2D gene mutation and the impact of its co-mutated genes on prognosis. Methods Clinical data of 155 newly diagnosed DLBCL patients were obtained. The second-generation sequencing method was used to detect 475 hotspot genes, including KMT2D mutation. Patients were divided into the KMT2D mutation group and KMT2D wild-type group based on the presence or absence of KMT2D gene mutation. Clinical characteristics, differences in co-mutated genes, and survival differences between the two groups were compared. Results The frequency of KMT2D mutation was 31%, which is predominantly observed in elderly patients (P=0.07) and less in the double-expressor phenotype (P=0.07). Compared with the KMT2D wild-type group, KMT2D gene mutation was associated with higher co-mutation rates of CDKN2A (OR=2.82, P=0.01) and BCL2 (OR=3.84, P=0.016), while being mutually exclusive with MYC gene mutation (OR=0.11, P=0.013). In univariate survival analysis, no statistically significant difference in overall survival (OS) was found between the KMT2D mutation group and the wild-type group (P=0.54). Further analysis of the prognostic significance of KMT2D with other gene mutations indicated that patients with KMT2D^mutBTG2^mut had poorer OS than those with KMT2D^wt BTG2^mut (P=0.07) and KMT2D^wt BTG2^wt (P=0.05). On the contrary, patients with KMT2D^mut CD79B^mut had better OS than those with KMT2D^mut CD79B^wt (P=0.09), with no prognostic impact observed for other co-mutated genes. Multivariate Cox regression analysis revealed that Ann Arbor stages Ⅲ and Ⅳ (HR=2.751, 95%CI: 1.169-6.472, P=0.02), elevated LDH levels (HR=2.461, 95%CI: 1.396-4.337, P=0.002), Ki-67 index>80% (HR=1.875, 95%CI: 1.066-3.299, P=0.029), and KMT2D^mutBTG2^mut(HR=4.566, 95%CI: 1.348-15.471, P=0.015) were independent risk factors for OS in patients with DLBCL (P<0.05). Conclusion DLBCL patients with KMT2D mutation often have multiple gene mutations, among which patients with a co-mutated BTG2 gene have poor prognosis.

Hydroxyectoine, a vital compatible solute, is widely utilized in cosmetics, food, pharmaceutical industries, and biologics. However, the current microbial fermentation methods for hydroxyectoine production face challenges including insufficient precursor supply and low yields. Therefore, developing engineering microbial strains capable of efficiently synthesizing hydroxyectoine is of great significance. In this study, we first constructed a high-yield ectoine-producing strain ECT04 by multi-copy integration of the ectoine synthesis genes ectABC into the pseudogene loci of Escherichia coli MG1655(DE3), achieving an ectoine titer of 6.03 g/L. Subsequently, we employed plasmids with varying copy numbers to express ectD from Chromohalobacter salexigens to enable the conversion for hydroxyectoine production. We further investigated the effects of promoter, co-substrate ɑ-ketoglutarate, Fe²⁺ concentration, and dissolved oxygen on hydroxyectoine synthesis. Through fed-batch fermentation in a 7-L bioreactor, we significantly enhanced the hydroxyectoine production efficiency, attaining a final titer of 8.58 g/L and a productivity of 0.24 g/(L·h). This work successfully achieved the de novo synthesis of hydroxyectoine in E. coli, laying a foundation for the efficient bioproduction of this compound.
Escherichia coli/genetics* ; Fermentation ; Amino Acids, Diamino/biosynthesis* ; Bioreactors/microbiology* ; Metabolic Engineering/methods* ; Chromohalobacter/genetics* ; Plasmids/genetics*

A solid-phase extraction-ultra-performance liquid chromatography-tandem mass spectrometry(SPE-UPLC-MS/MS)method was established for simultaneous determination of 10 kinds of neonicotinoid pesticide residues in aquaculture water.Based on the chemical properties of neonicotinoid pesticides and the matrix characteristics of aquaculture water,suitable temporary storage methods for water samples and appropriate solid-phase extraction columns were selected,and the extraction conditions(including elution solvents and sample loading volumes)were optimized.The method employed acetonitrile and 5 mmol/L ammonium acetate solution(containing 0.1%formic acid)as the mobile phase and an Oasis HLB solid-phase extraction column combined with PSA as a dispersive sorbent for sample purification.The method exhibited good linearity in detection of neonicotinoid in concentration range of 0.2-50 ng/mL(R2>0.99797),with a detection limit of 0.5 ng/L and a quantification limit of 1 ng/L,which were significantly lower than the maximum acceptable method detection limits(9-500 ng/L)for neonicotinoid insecticides in water published by the European Commission.In pond water,rice-fish water,and seawater,the average recoveries of the 10 target analytes were 74.6%-114.1%,with relative standard deviations ranging from 0.3%to 9.6%.Using this method,actual sample tests were conducted on the Pearl River water,Zhaoqing pond water,and Qingyuan rice-fish aquaculture water.The total concentration of five neonicotinoid pesticides in the Pearl River water ranged from 154.8 to 246.6 ng/L,the total concentration of four neonicotinoid pesticides in the Zhaoqing pond water was 95.0-176.1 ng/L,and the total concentration of three neonicotinoid pesticides in the Qingyuan rice-fish aquaculture water was 2.3-11.7 ng/L.This method was simple in operation,highly sensitive,and had strong resistance to interference.It was suitable for detection of trace neonicotinoid pesticides in aquaculture water and could provide technical support for construction of a green aquaculture environment and resolution of international trade disputes.

Objective To investigate the protective effect of dandelion flavone(DF)on lipopolysaccharide(LPS)-induced colon epithelial cell injury by intervening oxidative stress and inflammation with AT-specific binding protein 2(SATB2).Methods Colon epithelial cells FHC were cultured.FHC cells were randomly divided into control group(normal cultured),LPS group(10 μg·mL-1 LPS),experimental-L group(10 μg·mL-1 LPS+1 μmol·L-1 DF),experimental-H group(10 μg·mL-1 LPS+5 μmol·L-1 DF),experimental-H+sh-NC group(transfected with sh-NC+10 μg·mL-1 LPS+5 μmol·mL-1 DF),experimental-H+sh-SATB2 group(transfected with sh-SATB2+10 μg·mL-1 LPS+5μmol·L-1 DF).The relative expression level of SATB2 protein in FHC cells was detected by Western blotting.The survival rate of FHC cells in each group was determined by tetramethylazolium blue(MTT).The apoptosis rate of FHC cells in each group was detected by flow cytometry.The levels of malondialdehyde(MDA)and interleukin-6(IL-6)in FHC cells were detected by the kit.Results The relative expression levels of SATB2 protein in control group,LPS group,experimental-H group,experimental-H+sh-NC group and experimental-H+sh-SATB2 group were 0.83±0.09,0.19±0.03,0.66±0.05,0.62±0.07 and 0.23±0.03,respectively;cell viability rates were(100.00±1.00)％,(48.16±4.31)％,(85.31±5.83)％,(81.39±6.47)％and(58.75±5.24)％,respectively;cell apoptosis rates were(3.27±0.81)％,(41.26±2.09)％,(11.35±1.04)％,(10.29±1.26)％and(35.87±2.15)％,respectively;MDA levels were(13.16±1.73),(52.87±3.49),(23.19±2.05),(20.98±3.17)and(44.87±3.05)μmol·L-1,respectively;IL-6 levels were(507.18±103.26),(2 132.09±198.15),(883.16±136.92),(801.69±119.85)and(1 736.29±206.91)pg·mL-1,respectively.The above indicators in the LPS group showed significant differences compared to the control group(all P＜0.05);the above indicators in the experimental-H group showed significant differences compared to the LPS group(all P＜0.05);the above indicators in the experimental-H+sh-SATB2 group showed significant differences compared to the experimental-H+sh-NC group(all P＜0.05).Conclusion DF has a protective effect on LPS-induced colon epithelial cell injury by intervening oxidative stress and inflammation through SATB2.

Objective To explore the efficacy and safety of evolocumab in elderly patients with high-risk cardiovascular diseases.Methods A total of 153 patients with poor lipid control after conventional statin therapy who were hospitalized in the cardiologic departments in the First,Sec-ond,Sixth and Eighth Medical Centers of Chinese PLA General Hospital from November 2019 to November 2022 were included,and divided into non-elderly group(＜60 years old,46 cases),eld-erly group(60-74 years old,66 case)and very elderly group(≥75 years old,41 cases).They were all given evolocumab treatment according to guidelines.Another 50 over-75-year-old patients with high-risk cardiovascular diseases and poor lipid control who were hospitalized in the above cardiologic departments during the same period were treated with a statin drug combined with ezetimibe,and served as conventional treatment group(control group).The baseline clinical data and the blood indicators at 4th and 12th week after drug administration,and the occurrence of ad-verse drug reactions and major adverse cardiovascular events(MACE)within 12 weeks were com-pared among the groups.Results The levels of LDL-C and TC were significantly decreased in the three evolocumab treatment groups at 4 and 12 weeks after medication when compared with the baseline values(P＜0.05,P＜0.01),but there were no obvious differences in the 2 levels among the 3 groups at 12 weeks(P＞0.05).At the time point,no statistical difference was observed in the incidence of adverse events in the three groups(2.2％vs 3.0％vs 2.4％,P＞0.05).The levels of LDL-C and TC were decreased significantly in the very elderly group and the conventional treatment group at the 12th week when compared with the baseline levels(P＜0.05,P＜0.01),and the LDL-C level at the week was notably lower in the very elderly group than the convention-al treatment group(1.36±0.44 mmol/L vs 1.87±0.56 mmol/L,P＜0.01).But no difference was seen in the incidence of MACE between the 2 groups(12.2％vs 16.0％,P＞0.05),either in sur-vival rate between them(P=0.576).Conclusion For patients of all ages,evolocumab has good short-term efficacy in lipid control,and for those over 75 years old,the drug also shows good effi-cacy and sound safety.

AIM To study the Chemical constituents from Rhinacanthus nasutus(L.)Kurz and their in vitro antitumor and lipid-lowering activities.METHODS The ethyl acetate fraction from R.nasutus was isolated and purified by normal phase,reverse phase silica gel column and semi-preparative HPLC,then the structures of obtained compounds were identified by physicochemical properties and spectral data.The in vitro anti-tumor activity was determined by MTT assay,and the in vitro lipid-lowering activity was evaluated by oleic acid-induced HepG2 high-fat cell model.RESULTS Twenty compounds were isolated and identified as rhinacanthin A(1),rhinacanthin B(2),rhinacanthin C(3),rhinacanthin D(4),rhinacanthin E(5),rhinacanthin M(6),rhinacanthin N(7),rhinacanthin Q(8),rhinacanthin T(9),rhinacanthone(10),β-sitosterol(11),gallic acid(12),vanillic acid(13),syringic acid(14),lapachol(15),umbelliferone(16),sambucunlin A(17),17α,21-dihydroxy-1,4-pregnadiene-3,11,20-trione,21-acetate(18),1,8-dihydroxyanthraquinone(19),2,6-dimethoxy-1,4-benzoquinone(20).Compounds 3 and 7 inhibited the proliferation of HepG2,Hela,A549 and H22 tumor cells with IC50 values of(0.66±0.17)-(3.22±0.49)μmol/L in a time-and concentration-dependent manner.Compounds 3,7 and 9 had a lipid clearance rate of more than 50%in oleic acid-induced HepG2 high-fat cells.CONCLUSION Compounds 16-20 are isolated from this plant for the first time.Compounds 3 and 7 have good in vitro anti-tumor activities,and 3,7,9 have good in vitro lipid-lowering activities.

Objective:To compare the differences of clinical characteristics of temporal lobe epilepsy with bilateral hippocampal sclerosis (TLE-bHS) with those of temporal lobe epilepsy with unilateral hippocampal sclerosis (TLE-uHS).Methods:A retrospective analysis was performed. Forty-eight patients with confirmed TLE-bHS enrolled in Epilepsy Center, Department of Neurology, Second Affiliated Hospital, Medical School of Zhejiang University from January 2013 to January 2022 were chosen, and 101 patients with confirmed TLE-uHS admitted to our hospital at the same time period were selected as controls. Clinical data such as onset age, disease course, past medical history, seizure frequency, anti-seizure medications, video EEG and neuropsychological test results, and outcomes were analyzed.Results:Compared with the TLE-uHS group, the TLE-bHS group had higher male proportion, elder onset age, shorter disease course, higher seizure frequency, more types of past and currently used anti-seizure medications, lower proportion of autonomic nerve with aura, higher proportion of no aura at onset, higher proportion of slow head background movement in video EEG, and lower memory quotient, verbal memory scores and non-verbal memory scores, with significant differences ( P<0.05); the differences in ratio of past medical history and ratio of distributions of regions with interictal epileptiform abnormalities between the 2 groups were statistically significant ( P<0.05): the TLE-bHS group had significantly higher proportion of previous intracranial infection/encephalitis and higher ratio of bilateral temporal epileptiform abnormalities than the TLE-uHS group, while the TLE-uHS group had significantly higher proportion of patients with febrile convulsion history and higher ratio of unilateral temporal epileptiform abnormalities ( P<0.05). Only 10 patients (20.8%) in the TLE-bHS group received non-drug therapy, including anterior temporal lobectomy in 3 patients (Engel grading I in postoperative follow-up for 2 years), neuroregulatory therapy in 4, and ketogenic diet in 4; of the 55 patients (54.5%) in the TLE-uHS group who underwent anterior temporal lobectomy, 48 patients (87.3%) had Engel grading I, 1 patient (1.8%) had grading II, 4 (7.3%) had grading III, and 2 (3.6%) had grading IV after 2 years of follow-up. Conclusion:Differences in onset age, disease course, past medical history, seizure frequency, anti-seizure medications, and video EEG and neuropsychological test results can help to discriminate patients with TLE-bHS or with TLE-uHS.

Objective To investigate the activated phenotype and the expression of the receptor of advanced glycation endproducts(RAGE)of astrocytes after hypoxic-ischemic brain damage(HIBD)in neonatal rats and the effects of gastrodin(GAS)intervention on them.Methods Totally 48 neonatal 3 days SD rats were used to construct HIBD model and randomly divided into sham group,HIBD group and HIBD+GAS group(100 mg/kg),and the expressions of Al type astrocyte marker C3,A2 type astrocyte marker S100A10,RAGE,tumor necrosis factor-α(TNF-α),brain-derived neurotrophic factor(BDNF),and insulin-like growth factor(IGF-1)in the corpus callosum of the ischemic side were detected by Western blotting and immunohistochemical staining on day 1 and day 3 after HIBD.TNC-1 cells were divided into control group,oxygen glucose deprivation(OGD)group,OGD+GAS(0.34 mmol/L)group and GAS group,and then the protein expressions of RAGE,TNF-α,BDNF and IGF-1 were detected by Western blotting and immunofluorescence.Results In vivo,Western blotting showed that compared with the sham group,the protein expression levels of C3,S100A10,RAGE,TNF-α and IGF-1 in the 1 day and 3 days groups after HIBD group in 1 day group were significantly higher than those in the sham group(P＜0.05),but the protein expression level of BDNF decreased in 1 day group and increased in 3 days group(P＜0.05).Compared with the HIBD group,the C3,RAGE and TNF-α protein expression levels were significantly attenuated in the HIBD+GAS group(P＜0.05),and the protein expression levels of BDNF and IGF-1 further increased(P＜0.05).The protein expression of S100A10 in the 3 days group was higher than that in the HIBD group after GAS treatment(P＜0.05).The immunohistochemical staining results of C3,S100A10,and RAGE in the 1 day and 3 days groups after HIBD were consistent with Western blotting results.Furthermore,the protein expressions of RAGE and TNF-α were significantly enhanced in OGD-stimulated astrocytes(P＜0.05).After GAS intervention,while the expressions of both RAGE and TNF-α decreased significantly(P＜0.05),the expressions of BDNF and IGF-1 increased significantly(P＜0.05).Conclusion With inhibiting the up-regulation of RAGE signal in astrocyte after HIBD and expressions of A1 astrocyte and neuroinflammatory factors,gastrodin can promot the expressions of A2 astrocyte and nutritional factors,which play an important role in neuro-protective effect.

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.