Diterpenoid ferruginol is a key intermediate in biosynthesis of active ingredients such as tanshinone and carnosic acid.However, the traditional process of obtaining ferruginol from plants is often cumbersome and inefficient. In recent years, the increasingly developing gene editing technology has been gradually applied to the heterologous production of natural products, but the production of ferruginol in microbe is still very low, which has become an obstacle to the efficient biosynthesis of downstream chemicals, such as tanshinone. In this study, miltiradiene was produced by integrating the shortened diterpene synthase fusion protein,and the key genes in the MVA pathway were overexpressed to improve the yield of miltiradiene. Under the shake flask fermentation condition, the yield of miltiradiene reached about(113. 12±17. 4)mg·L~(-1). Subsequently, this study integrated the ferruginol synthase Sm CYP76AH1 and Sm CPR1 to reconstruct the ferruginol pathway and thereby realized the heterologous synthesis of ferruginol in Saccharomyces cerevisiae. The study selected the best ferruginol synthase(Il CYP76AH46) from different plants and optimized the expression of pathway genes through redox partner engineering to increase the yield of ferruginol. By increasing the copy number of diterpene synthase, CYP450, and CPR, the yield of ferruginol reached(370. 39± 21. 65) mg·L~(-1) in the shake flask, which was increased by 21. 57-fold compared with that when the initial ferruginol strain JMLT05 was used. Finally, 1 083. 51 mg·L~(-1) ferruginol was obtained by fed-batch fermentation, which is the highest yield of ferruginol from biosynthesis so far. This study provides not only research ideas for other metabolic engineering but also a platform for the construction of cell factories for downstream products.
Saccharomyces cerevisiae/genetics* ; Diterpenes/metabolism* ; Metabolic Engineering ; Fermentation ; Abietanes

Objective:To develop a robotic digestive endoscope system (RDES) and to evaluate its feasibility, safety and control performance by experiments.Methods:The RDES was designed based on the master-slave control system, which consisted of 3 parts: the integrated endoscope, including a knob and button robotic control system integrated with a gastroscope; the robotic mechanical arm system, including the base and arm, as well as the endoscopic advance-retreat control device (force-feedback function was designed) and the endoscopic axial rotation control device; the control console, including a master manipulator and an image monitor. The operator sit far away from the endoscope and controlled the master manipulator to bend the end of the endoscope and to control advance, retract and rotation of the endoscope. The air supply, water supply, suction, figure fixing and motion scaling switching was realized by pressing buttons on the master manipulator. In the endoscopy experiments performed on live pigs, 5 physicians each were in the beginner and advanced groups. Each operator operated RDES and traditional endoscope (2 weeks interval) to perform porcine gastroscopy 6 times, comparing the examination time. In the experiment of endoscopic circle drawing on the inner wall of the simulated stomach model, each operator in the two groups operated RDES 1∶1 motion scaling, 5∶1 motion scaling and ordinary endoscope to complete endoscopic circle drawing 6 times, comparing the completion time, accuracy (i.e. trajectory deviation) and workload.Results:RDES was operated normally with good force feedback function. All porcine in vivo gastroscopies were successful, without mucosal injury, bleeding or perforation. In beginner and advanced groups, the examination time of both RDES and ordinary endoscopy tended to decrease as the number of operations increased, but the decrease in time was greater for operating RDES than for operating ordinary endoscope (beginner group P=0.033; advanced group P=0.023). In the beginner group, the operators operating RDES with 1∶1 motion scaling or 5∶1 motion scaling to complete endoscopic circle drawing had shorter completion time [1.68 (1.40, 2.17) min, 1.73 (1.47, 2.37) min VS 4.13 (2.27, 5.16) min, H=32.506, P<0.001], better trajectory deviation (0.50±0.11 mm, 0.46±0.11 mm VS 0.82±0.26 mm, F=38.999, P<0.001], and less workload [42.00 (30.00, 50.33) points, 43.33 (35.33, 54.00) points VS 52.67 (48.67, 63.33) points, H=20.056, P<0.001] than operating ordinary endoscope. In the advanced group, the operators operating RDES with 1∶1 or 5∶1 motion scaling to complete endoscopic circle drawing had longer completion time than operating ordinary endoscope [1.72 (1.37, 2.53) min, 1.57 (1.25, 2.58) min VS 1.15 (0.86, 1.58) min, H=13.233, P=0.001], but trajectory deviation [0.47 (0.13, 0.57) mm, 0.44 (0.39, 0.58) mm VS 0.52 (0.42, 0.59) mm, H=3.202, P=0.202] and workload (44.62±21.77 points, 41.24±12.57 points VS 44.71±17.92 points, F=0.369, P=0.693) were not different from those of the ordinary endoscope. Conclusion:The RDES enables remote control, greatly reducing the endoscopists' workload. Additionally, it gives full play to the cooperative motion function of the large and small endoscopic knobs, making the control more flexible. Finally, it increases motion scaling switching function to make the control of endoscope more flexible and more accurate. It is also easy for beginners to learn and master, and can shorten the training period. So it can provide the possibility of remote endoscopic control and fully automated robotic endoscope.

Objective To investigate the clinical characteristics and prognosis of pneumococcal meningitis(PM),and drug sensitivity of Streptococcus pneumoniae(SP)isolates in Chinese children.Methods A retrospective analysis was conducted on clinical information,laboratory data,and microbiological data of 160 hospitalized children under 15 years old with PM from January 2019 to December 2020 in 33 tertiary hospitals across the country.Results Among the 160 children with PM,there were 103 males and 57 females.The age ranged from 15 days to 15 years,with 109 cases(68.1% )aged 3 months to under 3 years.SP strains were isolated from 95 cases(59.4% )in cerebrospinal fluid cultures and from 57 cases(35.6% )in blood cultures.The positive rates of SP detection by cerebrospinal fluid metagenomic next-generation sequencing and cerebrospinal fluid SP antigen testing were 40% (35/87)and 27% (21/78),respectively.Fifty-five cases(34.4% )had one or more risk factors for purulent meningitis,113 cases(70.6% )had one or more extra-cranial infectious foci,and 18 cases(11.3% )had underlying diseases.The most common clinical symptoms were fever(147 cases,91.9% ),followed by lethargy(98 cases,61.3% )and vomiting(61 cases,38.1% ).Sixty-nine cases(43.1% )experienced intracranial complications during hospitalization,with subdural effusion and/or empyema being the most common complication[43 cases(26.9% )],followed by hydrocephalus in 24 cases(15.0% ),brain abscess in 23 cases(14.4% ),and cerebral hemorrhage in 8 cases(5.0% ).Subdural effusion and/or empyema and hydrocephalus mainly occurred in children under 1 year old,with rates of 91% (39/43)and 83% (20/24),respectively.SP strains exhibited complete sensitivity to vancomycin(100% ,75/75),linezolid(100% ,56/56),and meropenem(100% ,6/6).High sensitivity rates were also observed for levofloxacin(81% ,22/27),moxifloxacin(82% ,14/17),rifampicin(96% ,25/26),and chloramphenicol(91% ,21/23).However,low sensitivity rates were found for penicillin(16% ,11/68)and clindamycin(6% ,1/17),and SP strains were completely resistant to erythromycin(100% ,31/31).The rates of discharge with cure and improvement were 22.5% (36/160)and 66.2% (106/160),respectively,while 18 cases(11.3% )had adverse outcomes.Conclusions Pediatric PM is more common in children aged 3 months to under 3 years.Intracranial complications are more frequently observed in children under 1 year old.Fever is the most common clinical manifestation of PM,and subdural effusion/emphysema and hydrocephalus are the most frequent complications.Non-culture detection methods for cerebrospinal fluid can improve pathogen detection rates.Adverse outcomes can be noted in more than 10% of PM cases.SP strains are high sensitivity to vancomycin,linezolid,meropenem,levofloxacin,moxifloxacin,rifampicin,and chloramphenicol.[Chinese Journal of Contemporary Pediatrics,2024,26(2):131-138]

Objective:To investigate the accuracy of next-generation sequencing technology(NGS)in detecting the polymorphisms of HLA-DRB1,DQB1,DQA1,DRB3,DRB4,DRB5,DPA1 and DPB1 alleles in randomly-selected unrelated healthy individuals from Shenzhen Han population,investigate the potential reason for HLA-DRB1 allele dropout in routine NGS,and establish an internal quality control system.Methods:NGS-based HLA class Ⅱ genotyping was performed on 1 012 samples using the MiSeqDxTM platform.The suspected missed alleles indicated by the quality control software and HLA-DRB1 homozygotes were confirmed by PCR-SSOP or PCR-SBT methods.Results:A total of 139 alleles were detected,including HLA-DRB1(45),DRB3(7),DRB4(5),DRB5(7),DQA1(17),DQB1(21),DPA1(10)and DPB1(27).HLA-DRB 1*09:01(17.09％),15:01(10.72％);DRB3*02:02(25.99％),03:01(10.18％);DRB4*01:03(36.46％);DRB5*01:01(15.42％);DQA1*01:02(20.01％),03:02(17.19％);DQB1*03:01(19.47％),03:03(17.98％),05:02(11.66％),06:01(10.67％);DPA1*02:02(54.45％),01:03(31.18％)and DPB1*05:01(39.13％),02:01(16.90％)alleles were the most common alleles in Shenzhen Han population(frequencies＞10％).There was no statistical difference between the gene frequencies of HLA-DRB1 and DQB1 loci in our study.The HLA Common and Well-Documented Alleles in China(CWD2.4)(x2=12.68,P＞0.05).94 cases of HLA-DRB1 homozygous samples detected by NGS were retested by PCR-SSOP or SBT method,and one case of allele dropout at HLA-DRB1 locus was found.SBT method confirmed that the allele of DRB1*04:03 was missed.The laboratory internal quality control system was established.Two cases of new alleles were detected and named by WHO Nomenclature Committee for Factors of the HLA System.Conclusion:The HLA genotyping results based on NGS showed a significantly lower ambiguity rate.The HLA class Ⅱ alleles exhibit genetic polymorphism in the Han population of unrelated healthy individuals in Shenzhen.The independent method based on NGS in clinical histocompatibility testing has limitations and requires internal quality control strategies to avoid allele-dropout events.

The paper summarizes the definition, location and main indication of qicimai points recorded in Huangdi Neijing (Yellow Emperor 's Inner Classic). It is found that qicimai points are the "upward moving points" in reference to the meridian distribution rule of "rooting, running, infusing and moving"; and corresponding to the sites of "running outwards and inwards" of the meridians' "separating, meeting and running outwards and inwards". It also includes the infusing points for the sea of qi and marrow. The new idea, "selecting qicimai points for the treatment of qi obstruction in the neck gate", is proposed. Based on the systematic application of the acupoints on the nape region, it is anticipated that a new approach will be provided to the treatment of the diseases in the neck, shoulder, head, face, the five sensory organs, mental disorders and zangfu qi dysfunction.
Humans ; Meridians ; Acupuncture Points ; Acupuncture Therapy

OBJECTIVE: To observe the effects of electroacupuncture (EA) on cardiac function and local field potential (LFP) in sensory and motor cortices in mice with stress cardiomyopathy (SC), and to explore the possible mechanism of EA in improving SC. METHODS: Twenty-seven female C57BL/6 mice were randomized into a blank group, a model group and an EA group, 9 mice in each group. In the model group and the EA group, SC model was established by continuous intraperitoneal injection of isoproterenol (ISO) for 14 days. At the same time of modeling, EA was applied at "Neiguan" (PC 6) and "Shenmen" (HT 7) in the EA group, with disperse-dense wave, in frequency of 2 Hz/15 Hz, 15 min each time, once a day for 14 days. After intervention, the total movement distance, the number of crossing grid and the number of crossing central grid of open field test within 5 minutes were observed; the left ventricular function indexes (left ventricular diameter of end-diastole [LVIDd], left ventricular diameter of end-systole [LVIDs], left ventricular volume of end-diastole [LVEDV], left ventricular volume of end-systole [LVESV], ejection fraction [EF] and fraction shortening [FS]) were detected by echocardiography; the changes in ST-segment amplitude and PR interval of electrocardiogram were observed; the morphology of myocardial tissue was observed by HE staining; the serum levels of cortisol (CORT), cardiac troponin T (cTnT) and brain natriuretic peptide (BNP) were detected by ELISA; the changes of LFP in sensory and motor cortices were recorded by Plexon multi-channel acquisition system. RESULTS: Compared with the blank group, in the model group, the total movement distance, the number of crossing grid and the number of crossing central grid of open field test were decreased (P<0.05); LVIDd, LVIDs, LVEDV and LVESV were increased (P<0.05), EF and FS were decreased (P<0.05); ST-segment amplitude was increased (P<0.05) and PR interval was prolonged (P<0.05); irregular myocardial fiber arrangement, interstitial edema and inflammatory cell infiltration were observed; the serum levels of CORT, cTnT and BNP were increased (P<0.05); in the sensory cortex, the ratios of delta, theta, alpha and beta frequency bands were increased (P<0.05), the maximum energy spectrum of theta and beta frequency bands was increased (P<0.05), the power spectral density (PSD) of delta, theta, alpha, beta and gamma frequency bands was increased (P<0.05); in the motor cortex, the ratios of delta, theta, alpha and beta frequency bands were increased (P<0.05), the maximum energy spectrum as well as PSD of delta, theta, alpha, beta and gamma frequency bands were increased (P<0.05). Compared with model group, in the EA group, the total movement distance, the number of crossing grid and the number of crossing central grid of open field test were increased (P<0.05); LVIDd, LVIDs, LVEDV and LVESV were decreased (P<0.05), EF and FS were increased (P<0.05); ST-segment amplitude was decreased (P<0.05), and the PR interval was shortened (P<0.05); myocardial fiber injury and inflammatory cell infiltration were reduced; the serum levels of CORT, cTnT and BNP were decreased (P<0.05); in the sensory cortex, the ratios of theta, alpha and beta frequency bands were decreased (P<0.05), the ratio of gamma frequency band was increased (P<0.05), the maximum energy spectrum of theta frequency band as well as the PSD of theta, alpha, beta and gamma frequency bands were decreased (P<0.05); in the motor cortex, the ratios of theta, alpha and beta frequency bands were decreased (P<0.05) and the ratio of gamma frequency band was increased (P<0.05), the maximum energy spectrum of delta frequency band was increased (P<0.05), the maximum energy spectrum of theta frequency band as well as the PSD of theta and gamma frequency bands were decreased (P<0.05). CONCLUSION EA can improve cardiac function in mice with stress cardiomyopathy, and its mechanism may be related to the regulation of local field potentials in sensory and motor cortices.
Female ; Mice ; Animals ; Electroacupuncture ; Takotsubo Cardiomyopathy ; Motor Cortex ; Mice, Inbred C57BL ; Myocardium

This study aimed to investigate the development status of traditional Chinese medicine(TCM) intervention in psoriasis in recent ten years, analyze the research hotspots, and summarize the development trends to provide reference materials for scholars in this field. Taking the available literature related to the field of TCM intervention in psoriasis as the research object, the trends, contents, and source publications were statistically analyzed based on bibliometrics. The research cooperation and co-occurrence of keywords in this field were studied by the knowledge map analysis method based on CiteSpace. The total number of Chinese papers was 2 993 and English papers 285. In terms of publication trend, the annual publication of English papers was low but showed an obvious upward trend, while the increase in Chinese papers fluctuated and tended to be flat. In terms of the content of Chinese papers published, TCM ranked first according to the discipline(2 415). In English papers, the number of publications in pharmacology and pharmaceutical science was the highest(87). Literature source analysis showed that the Chinese and English journals with the most publications were China Journal of Traditional Chinese Medicine and Pharmacy and Evidence Based Complementary and Alternative Medicine, respectively. Beijing University of Chinese Medicine published the most dissertations in China(99). The authors with the most publications in Chinese and English were LI Bin(Yueyang Hospital of Integrated Traditional Chinese and Western Medicine Affiliated to Shanghai University of Traditional Chinese Medicine) and LU Chuan-jian(Guangdong Hospital of Traditional Chinese Medicine). As revealed by the CiteSpace analysis of the research cooperation network, there were four mature and stable core teams in this field, but the cooperation intensity between different teams was weak. According to the keywords co-occurrence knowledge graph constructed by CiteSpace, the current hot keywords in this field are as follows: psoriasis, blood-heat syndrome, blood-stasis syndrome, fire needle, blood-dryness type, imiquimod, TCM bath, etiology and pathogenesis, cytokines, cupping therapy, etc. In summary, Chinese scholars have conducted active exploration and research in the field of TCM intervention in psoriasis in recent ten years. The overall development trend is good, and the breadth and depth of the research are constantly extending. It is suggested that relevant research should be free from discipline restrictions and strive for interdisciplinary integration.
Humans ; Medicine, Chinese Traditional ; Psoriasis/drug therapy*

Chronic inflammation induced by Helicobacter pylori is considered to be one of the main causes of gastric cancer,and CagA is a main virulence factor of H.pylori.The study aimed to investigate the role and mechanism of CagA in host inflammatory response.Mass spectrometry was used to identify the interacting proteins of CagA in AGS cells.By immunoprecipitation and immunofluorescence,the interaction was validated.Pathway expression was detected by immunoblotting after knockdown by using siRNA,and mRNA levels of inflammatory cytokines were detected by quantitative PCR.CagA-induced inflammatory responses were detected in clinical samples using hemoglobin-eosin staining(H&E).Data showed that CagA interacted with SHARPIN.And CagA activated the NF-κB signaling pathway and upregulated the mRNA and protein levels of the inflammatory cytokines IL-6,IL-8,and TNF-α,as compared with the CagA knockout strain(all P<0.05).Knockdown of SHARPIN by siRNA reduced inflammation levels and partially inhibit NF-κB signaling.In clinical samples,CagA-positive samples exhibited stronger inflammatory responses.To sum up,CagA promoted the host inflammatory response,and CagA-induced inflammatory response was reduced when SHARPIN was partially inhibited,suggesting that CagA activates the NF-κB signaling pathway through binding to SHARPIN.

OBJECTIVE: Prenatal phthalate exposure has been associated with placental inflammatory factors and infant allergic rhinitis (AR). However, the results are inconclusive. We designed a population-based cohort study to examine the effects of placental inflammatory biomarkers on the sex-dependent associations between maternal phthalate exposure and infant AR. METHODS: A total of 2,348 pregnant women from Ma'anshan, Anhui Province, China, who were screened before antenatal visits and met the inclusion criteria, were included in the present study. We assessed AR in their offspring aged 36 months with a questionnaire. Quantitative PCR was performed to measure placental inflammatory factor mRNAs. The independent samples t-test and multivariable logistic regression were used to determine the associations between infant AR and maternal phthalates. RESULTS: Childhood AR may be related to education and family monthly income ( P = 0.01). The phthalate metabolites, mono (2-ethylhexyl) phthalate (MEHP), mono (2-ethyl-5-hydroxyl) phthalate (MEHHP), in pregnant women were associated with a significantly increased risk for infant AR in males [ P < 0.05; odds ratio ( OR): 1.285; 95% confidence interval ( CI): 1.037-1.591, and OR: 1.232, 95% CI: 1.008-1.507, respectively], but not females. Additionally, irritably-increased expression levels of HO-1 and IL-4 were associated with AR in male infants ( OR: 1.175; 95% CI: 1.038-1.329 and OR: 1.181; 95% CI: 1.056-1.322, respectively). The association between maternal urinary MEHHP and placental HO-1 was marginally significant according to mediation analysis. CONCLUSION The associations of maternal MEHHP and MEOHP levels with fetal AR in males were significant. Placental HO-1 was a fractional mediator in the associations between MEHHP and AR. Thus, the placenta should be further investigated as a potential mediator of maternal exposure-induced disease risk in children.
Biomarkers ; Child, Preschool ; Cohort Studies ; Diethylhexyl Phthalate/analogs & derivatives* ; Female ; Humans ; Interleukin-4/pharmacology* ; Male ; Maternal Exposure/adverse effects* ; Phthalic Acids/adverse effects* ; Placenta ; Pregnancy ; Rhinitis, Allergic/epidemiology*

Objective: To explore the heterogeneity and correlation of clinical phenotypes and genotypes in children with disorders of sex development (DSD). Methods: A retrospective study of 1 235 patients with clinically proposed DSD in 36 pediatric medical institutions across the country from January 2017 to May 2021. After capturing 277 DSD-related candidate genes, second-generation sequencing was performed to analyzed the heterogeneity and correlation combined with clinical phenotypes. Results: Among 1 235 children with clinically proposed DSD, 980 were males and 255 were females of social gender at the time of initial diagnosis with the age ranged from 1 day of age to 17.92 years. A total of 443 children with pathogenic variants were detected through molecular genetic studies, with a positive detection rate of 35.9%. The most common clinical phenotypes were micropenis (455 cases), hypospadias (321 cases), and cryptorchidism (172 cases) and common mutations detected were in SRD5A2 gene (80 cases), AR gene (53 cases) and CYP21A2 gene (44 cases). Among them, the SRD5A2 mutation is the most common in children with simple micropenis and simple hypospadias, while the AMH mutation is the most common in children with simple cryptorchidism. Conclusions: The SRD5A2 mutation is the most common genetic variant in Chinese children with DSD, and micropenis, cryptorchidism, and hypospadias are the most common clinical phenotypes. Molecular diagnosis can provide clues about the biological basis of DSD, and can also guide clinicians to perform specific clinical examinations. Target sequence capture probes and next-generation sequencing technology can provide effective and economical genetic diagnosis for children with DSD.
3-Oxo-5-alpha-Steroid 4-Dehydrogenase/genetics* ; Child ; China/epidemiology* ; Cryptorchidism/genetics* ; Disorders of Sex Development/genetics* ; Female ; Genital Diseases, Male ; Genotype ; Humans ; Hypospadias/genetics* ; Male ; Membrane Proteins/genetics* ; Penis/abnormalities* ; Phenotype ; Retrospective Studies ; Steroid 21-Hydroxylase/genetics*