Normalized spectral settings in light- and myopia-related experimental studies / 中华实验眼科杂志

There are more and more studies investigating the influence of spectral difference on myopia, but the settings of light sources are different in each study.As a physical quantity, light can be measured radiometrically, mainly to detect the energy properties of light.As a psychological and physical quantity, the light should be measured photometrically to detect the degree of light perception of the organism.Different optical measuring tools measure based on the response curve of the photoelectric conversion device.The essence of the light source spectrum setting in the experiment is to control the sensitivity of organisms to light.Therefore, special attention should be paid to the influence of light source setting differences when interpreting similar experimental results.When setting the light in animal experiments, it is necessary to consider the differences in the absorption curve of the animal's photosensitive pigment and the curve of the animal's spectral light efficiency.It is also suggested that a radiometer should be used to unify the photon flux density consistent among different groups based on 200-300 lx light intensity.In this paper, the normalization of spectral settings in experiments related to light and myopia was reviewed.

Comparison of the effects of different chromatic lights on accommodative response and microfluctuation in myopes and emmetropes / 中华实验眼科杂志

Objective:To explore the effects of conflicting stimuli generated by different chromatic lights on visual display terminal (VDT) on accommodative response and microfluctuation of myopes and emmetropes, and to investigate the possible relationship between chromatic light, accommodation and the development and progression of myopia.Methods:A non-randomized controlled trial was conducted.Forty-one subjects aged 22 to 30 years old were enrolled, including 19 emmetropes in emmetropic group and 22 myopes in myopic group.The subjects had the normal color vision and no ocular organic diseases.The interventions were screens of different colors.There were 7 chromatic light conditions, including 3 monochromatic lights (red, green, blue), 3 bichromatic lights (red+ green, red+ blue, green+ blue) and 1 polychromatic light (white=red+ green+ blue). Subjects were asked to look at a black E target on a VDT at a distance of 33 cm for more than 20 seconds.The background color of the VDT was changed randomly in the 7 chromatic light conditions.The accommodative responses were recorded with the Grand Seiko WAM-5500 automatic infrared refractor every 0.2 seconds and the accommodative microfluctuation was calculated as the standard deviation of the accommodative response.Accommodative response and accommodative microfluctuation under different chromatic light conditions were compared.This study adhered to the Declaration of Helsinki.The study protocol was approved by the Ethics Committee of the First Affiliated Hospital, Zhejiang University School of Medicine (No.2019-1564). Written informed consent was obtained from each subject.Results:No statistically significant difference was found in the accommodative response between the two groups ( Fgroup=2.626, P=0.113). There was a statistically significant difference under different chromatic light conditions between the two groups ( Flight=39.070, P<0.01). There were similar trends in the effects of various color lights in both groups, with the largest accommodative response under monochromatic red light, followed by the bichromatic light containing red light, and then the smallest accommodative response under monochromatic blue light, and the differences were statistically significant (all at P<0.05). The accommodative microfluctuations under red, green, blue, red+ blue, red+ green, blue+ green and white light conditions were (0.142±0.033), (0.128±0.038), (0.131±0.043), (0.139±0.039), (0.127±0.034), (0.131±0.043) and (0.139±0.042)D in emmetropic group, and (0.178±0.043), (0.164±0.043), (0.159±0.039), (0.174±0.042), (0.166±0.036), (0.159±0.031) and (0.174±0.035)D in myopic group, respectively, showing statistically significant differences between them ( Fgroup=12.146, P<0.01; Flight=2.782, P<0.05). The accommodative microfluctuations under the 7 light conditions were higher in myopic group than in emmetropic group, and the differences were statistically significant (all at P<0.05). In myopes, the accommodative microfluctuation was the largest under red light, which was significantly larger than that under blue light, and was the smallest under blue+ green light (all at P<0.05). There was no significant difference in the accommodative microfluctuation between bichromatic light and its two monochromatic lights, or between the polychromatic light (white light) and its three monochromatic lights (all at P>0.05). There was no significant effect of various chromatic lights on the accommodative microfluctuation in emmetropic group (all at P>0.05). Conclusions:The accommodative microfluctuation is greater in myopes than in emmetropes.The stimuli produced by long-wavelength light cause larger accommodative microfluctuation, while conflicting stimuli generated by different chromatic lights do not increase accommodative microfluctuation.

Cloning and expression analysis of superoxide dismutase SmMSD2 gene from Salvia miltiorrhiza / 药学学报

Superoxide dismutase (SOD) is a key enzyme that scavenge superoxide anion free radical (O2·-) in vivo, and plays an important role in plant growth and development and stress. In this study, according to the genome and transcriptome data of Salvia miltiorrhizae, 9 SOD genes were identified and the expression patterns of SOD family genes were further analyzed, including 5 Cu/Zn-SOD, 2 Fe-SOD and 2 Mn-SOD. On the basis of proteomic analysis, combined with transcriptome data, one full-length cDNA of Mn-SOD gene, namely SmMSD2 was cloned from Salvia miltiorrhizae. The results of amino acid sequence alignment and phylogenetic analysis showed that SmMSD2 protein belongs to the manganese superoxide dismutase (Mn-SOD) subfamily, and SmMSD2 protein shares high sequence identity with the Mn-SOD proteins of various plants that all contain a C-terminal conserved metal-binding domain "DVWEHAYY". The prokaryotic expression vector pMAL-c2X-SmMSD2 was constructed and transformed into E. coli BL21 expressing strain, and the target recombinant protein was successfully induced and its enzymatic properties were analyzed. Spatiotemporal expression analysis showed that SmMSD2 gene was expressed in all tissues, indicating that SmMSD2 gene was constitutively expressed at a stable level. Real-time quantitative PCR indicated that drought (15% PEG6000), abscisic acid (ABA) and indole-3-acetic acid (IAA) could induce the expression of SmMSD2 gene, suggesting that SmMSD2 may be involved in the response of Salvia miltiorrhizae to abiotic stress such as drought, as well as the signaling pathways of phytohormone ABA and IAA. These results lay the foundation for further elucidating the involvement of superoxide dismutase in the stress response and accumulation of active components of Salvia miltiorrhiza.

Systematic identification of chemical forms of key terpene synthase in Cinnamomum camphora / 中国中药杂志

Cinnamomum camphora is an important economic tree species in China. According to the type and content of main components in the volatile oil of leaf, C. camphora were divided into five chemotypes, including borneol-type, camphor-type, linalool-type, cineole-type, and nerolidol-type. Terpene synthase(TPS) is the key enzyme for the formation of these compounds. Although several key enzyme genes have been identified, the biosynthetic pathway of(+)-borneol, which has the most economic value, has not been reported. In this study, nine terpenoid synthase genes CcTPS1-CcTPS9 were cloned through transcriptome analysis of four chemical-type leaves. After the recombinant protein was induced by Escherichia coli, geranyl pyrophosphate(GPP) and farnesyl pyrophosphate(FPP) were used as substrates for enzymatic reaction, respectively. Both CcTPS1 and CcTPS9 could catalyze GPP to produce bornyl pyrophosphate, which could be hydrolyzed by phosphohydrolase to obtain(+)-borneol, and the product of(+)-borneol accounted for 0.4% and 89.3%, respectively. Both CcTPS3 and CcTPS6 could catalyze GPP to generate a single product linalool, and CcTPS6 could also react with FPP to generate nerolidol. CcTPS8 reacted with GPP to produce 1,8-cineol(30.71%). Nine terpene synthases produced 9 monoterpene and 6 sesquiterpenes. The study has identified the key enzyme genes responsible for borneol biosynthesis in C. camphora for the first time, laying a foundation for further elucidating the molecular mechanism of chemical type formation and cultivating new varieties of borneol with high yield by using bioengineering technology.

Serum levels of megaprolactin and gonadal hormones in patients with hyperprolactinemia induced by antipsychotics and their clinical significance / 中国基层医药

Objective:To investigate serum levels of macroprolactin and gonadal hormones in patients with hyperprolactinemia induced by antipsychotics and their clinical significance.Methods:A total of 105 female patients with schizophrenia who received treatment in Huzhou Third Municipal Hospital from June 2017 to October 2018 were included in this study. All these patients received the antipsychotic drug clozapine for 2 months. Then 50 patients with hyperprolactinemia were included in the observation group, and 55 patients who had no hyperprolactinemia were included in this control group. The scores of the Scale for the Assessment of Negative Symptoms and Scales for the Assessment of Positive Symptoms were compared between the two groups. Serum levels of macroprolactin, progesterone, testosterone, estradiol, prolactin, luteinizing hormone, and follicle-stimulating hormone were then compared between the two groups. The Spearman correlation analysis was used to analyze the correlation between serum macroprolactin level and serum progesterone, estradiol, prolactin, and luteinizing hormone levels.Results:Serum macroprolactin level in the observation group was significantly higher than that in the control group [(63.80 ± 12.13) ng/mL vs. (59.07 ± 9.84) ng/mL, t = 2.20, P = 0.030). Serum levels of progesterone, testosterone, estradiol, prolactin, luteinizing hormone, and follicle-stimulating hormone were (4.01 ± 0.47) ng/mL, (5.59 ± 1.15) ng/mL, (236.72 ± 15.14) pg/mL, (127.30 ± 12.40) ng/mL, (6.05 ± 1.10) mIU/mL, (8.52 ± 2.13) mIU/mL, respectively, and they were (10.25 ± 1.83) ng/mL, (6.01 ± 1.20) ng/mL, (433.10 ± 20.90) pg/mL, (50.58 ± 6.22) ng/mL, (7.69 ± 2.36) mIU/mL, (8.48 ± 2.01) mIU/mL, respectively in the control group. Serum levels of progesterone, estradiol, and luteinizing hormone in the observation group were significantly lower than those in the control group, and serum level of prolactin in the observation group was significantly higher than that in the control group ( t = 23.41, 54.66, 4.63, 40.61, all P < 0.05). There were no significant differences in serum levels of testosterone and follicle-stimulating hormone between the two groups ( t = 1.82, 0.09, P = 0.071, 0.921). Spearman correlation analysis results showed that serum macroprolactin level was negatively correlated with serum levels of progesterone and estradiol, and it was positively correlated with serum levels of prolactin and luteinizing hormone ( r = -0.42, -0.51, -0.68, 0.70, all P < 0.05). Conclusion:Serum levels of macroprolactin and prolactin were higher, and serum levels of progesterone, estradiol, and luteinizing hormone levels were lower in patients with hyperprolactinemia induced by antipsychotics than in patients without hyperprolactinemia. Serum levels of macroprolactin, prolactin, luteinizing hormone, progesterone, and estradiol were remarkably correlated with the balance of gonadal hormones. The study outcomes are of great innovation and science.

Cloning, induction pattern and prokaryotic expression of a small heat shock protein SmHSP21.8 gene from Salvia miltiorrhiza / 药学学报

In order to reveal the molecular mechanism of the small heat shock proteins (sHSPs) involved in stress resistance and active ingredients accumulation in Salvia miltiorrhiza, a small heat shock protein gene was cloned from Salvia miltiorrhiza by reverse transcription PCR according to the transcriptome data of orange root Salvia miltiorrhiza. The gene is named SmHSP21.8 based on the molecular weight of the protein, and it contains an open reading frame of 585 bp, which encodes 194 amino acids. The results of phylogenetic analysis and amino acid sequence alignment showed that SmHSP21.8 protein belongs to the endoplasmic reticulum (ER) subfamily, and contains a conserved endoplasmic reticulum-specific DPFR-I/V-LE-H/Q-x-P motif at N-terminus. The prokaryotic expression vector pMAL-c2X-SmHSP21.8 was constructed and transformed into E. coli BL21 competent cells. The recombinant protein was successfully expressed after inducted. Temporal and spatial expression analysis showed that SmHSP21.8 gene was the highest expressed in flowers and had significant tissue specificity. The relative expression of the gene was significantly increased in seedlings after induction by 38 ℃, PEG6000, abscisic acid(ABA), and indole-3-acetic acid (IAA), indicating that SmHSP21.8 gene may be involved in abiotic stress such as high temperature and drought, as well as the response to exogenous hormones ABA and IAA. These results lay the foundation for further research on the molecular mechanism of small heat shock proteins involved in adversity stress.

Daily violence exposure and its impact on campus bullying among middle school students in Xinxiang / 中国学校卫生

Objective@#The study is to explore daily violence exposure and its association with campus bullying, to provide theoretical basis for positive development of middle school students.@*Methods@#Questionnaire survey was conducted by using Violence Exposure Scale, Normative Beliefs about Aggression Scale, Middle School Students’ Self-control Scale, and Middle School Students’ Campus Bullying Scale. During Aug. to Oct. 2019, 1 372 middle school students were selected by the convenient sampling method as subjects of study from 2 junior high schools and 3senior high schools in Xinxiang.@*Results@#The total score in daily violence exposure was (34.22±12.09). The scores of violence exposure, traditional bullying and cyberbullying in female were lower than in male(t=-2.60--6.32, P<0.05). The scores of violence exposure, traditional bullying and cyberbullying in junior high school students were higher than senior high school students(t=4.59-7.50, P<0.05). The relationship between violence exposure and normative beliefs about aggression, traditional bullying, cyberbullying were positive (r=0.20, 0.44, 0.51, P<0.01). The relationship between violence exposure and self-control was negative (r=-0.29, P<0.01) . The relationship between normative beliefs about aggression and traditional bullying, cyberbullying were positive (r=0.28, 0.22, P<0.01). The relationship between normative beliefs about aggression and self-control was negative (r=-0.38, P<0.01). Violence exposure indirectly affects traditional bullying/cyberbullying through normative beliefs about aggression. The effect of normative beliefs about aggression on the traditional bullying/cyberbullying of middle school students is reduced with the increase of self-control.@*Conclusion@#Normative beliefs about aggression plays an intermediary role in violence exposure and traditional bully/cyberbullying, and self-control regulates the relationship between them.

Cloning and expression analysis of 5-phosphomevalonate kinase gene (CcPMK) in Cinnamomum camphora / 中国中药杂志

The 5-phosphomevalonate kinase(PMK) is a key enzyme in mevalonate(MVA) pathway which reversibly catalyzes the phosphorylation of mevalonate 5-phosphate(MVAP) to form mevalonate-5-diphosphate(MVAPP) in the presence of ATP and divalent metal ion such as Mg~(2+). In this research, on the basis of the transciptome database of Cinnamomum camphora, the PMK was cloned by cDNA from C. camphora, and was named CcPMK(GenBank number KU886266). The ORF of CcPMK was composed of 1 545 bp, encoding 514 amino acids. The bioinformatics analysis of CcPMK indicated that the molecular weight of the encoded protein was 56.14 kDa, with a theoretically isoelectric point of 7.64, and there was no signal peptide and transmembrane structure in putative protein. By multiple sequence alignment and phylogenetic tree analysis, we found that similarity between CcPMK and PMK amino acid sequence of other plants was as high as 75%. Among the similar sequences, 45% of them belonged to the alpha helix, while 16% belonged to the beta strand. CcPMK obtained 3 PMK protein family motifs and 1 ATP binding site Gly-Leu-Gly-Ser-Ser-Ala-Ala, and its 3 D structure contained a catalytic pocket structure, proving CcPMK as a member of PMK gene family. The result of phylogenetic tree showed that CcPMK was closely related to monocotyledon plants such as Phonenix dactylifera. The results of the Real-time PCR indicated that the expression level of CcPMK in borneol type was higher than that in linalool type, cineol type, iso-nerolidol type and camphor type. CcPMK expressed highest in roots and lowest in branches. Our results revealed that the expression level of CcPMK was different among five chemical types and different plant tissues, and the research provides foundation for further study of the terpenoids biosynthetic pathway in C. camphora.

Cloning and expression analysis of 3-hydroxy-3-methylglutaryl coenzyme A reductase (CcHMGR) genes in Cinnamomum camphora (L.) Presl / 药学学报

3-Hydroxy-3-methylglutaryl coenzyme A reductase (HMGR) is the first rate-limiting enzyme of terpenoid biosynthesis in the mevalonic acid pathway (MVA) pathway. It is an important regulatory site in terpenoids metabolism pathway in the cytoplasm. According to the transcriptome database of Cinnamomum camphora, two HMGRs named CcHMGR1 (GenBank: MN163055) and CcHMGR2 (GenBank: MN163056) were cloned by cDNA from C. camphora. The ORF of CcHMGR1 and CcHMGR2 is composed of 1 689 bp and 1 683 bp, respectively, encoding 562 and 560 amino acids. The bioinformatics analysis of CcHMGR1 and CcHMGR2 indicated that the molecular weight of the encoded protein is 59.819 kDa and 59.397 kDa, with a theoretically isoelectric point of 8.20 and 8.61, respectively. There are 2 transmembrane structures without signal peptide existing in the encoded amino acid of CcHMGRs. The analysis of sequence alignment and phylogenetic tree showed that theCcHMGRs belonged to the HMGR family. The camphor is divided into five chemitypes, according to the main chemical compoundsin C. camphora. The results of the real time PCR indicated that the expression level of CcHMGRs in Cineol type was higher than that in Linalool type, iso-nerolidol type, Camphor type and Borneol type. CcHMGRs expressed highest in roots and lowest in branches. In this study, the cDNA full length of CcHMGRs were cloned from C. camphora for the first time. Our results revealed that the expression level of CcHMGRs were different among five chemical types and different plant tissues, and the research provides foundation for further study of the terpenoids biosynthetic pathway in C. camphora.

Discovery and Application of Quality Marker of Traditional Chinese Medicine Based on Integrative Pharmacology / 中国实验方剂学杂志

Quality marker(Q-marker) is a new concept and pattern for quality control of traditional Chinese medicine(TCM),which will lead the development direction for quality control of TCM.Among them,how to characterize the overall quality attribute of TCM and its biological effect,is a critical scientific problem in the study of Q-marker.In this paper,integrated pharmacology is utilized to screen out and confirm the Q-marker from the complex system of TCM,so as to solve the critical scientific problem.System biology in vivo is firstly applied to establish the correlation of chemical fingerprints of TCM,their metabolic fingerprints,network targets,biological effects and efficacy of TCM,which is used to preliminary screen out Q-marker of TCM.Following that,a pharmacological method in vitro,including intestinal absorption in vitro coupled with bioactivity assessment,is employed to simultaneously determine the absorbed doses of TCM and evaluate their biological activity.Furthermore,data mining is utilized to establish the exact quantitative mathematic model between Q-marker of TCM and bioactivity.Meanwhile,two representative examples,including Yuanhu Zhitong tablets,Xinsuning capsules,are introduced to identify Q-marker of TCM and establish their quality standards related with bioactivity,which will be beneficial to improve the level of quality control of TCM and ensure the effectiveness and safety of clinical applications.

Predictive value of in vitro methyl thiazolyl tetrazolium assay chemosensitivity test of cytotoxic drug activity in breast cancer / 中国综合临床

Objective To investigate the value of methyl thiazolyl tetrazolium assay ( MTT) in predicting drug sensitivity of breast cancer cells in vitro. Methods From January 2010 to July 2016,one hundred and ninety-two patients with breast cancer who underwent modified radical mastectomy or breast conserving surgery (no preoperative radiotherapy or chemotherapy) in the Shanghai Fengxian District Central Hospital were selected. MTT method was used to determine the inhibitory level and sensitivity of 12 drugs and 3 chemotherapy regimens to primary cultured cancer cells of 192 patients with breast cancer. Results (1) The sensitivity of breast cancer cells to 12 drugs were in sequence from high to low as follows: Paclitaxel (PTX)＞ Epirubicin ( EPI )＞ Cisplatin ( DDP )＞ 5-Fluorouracil ( 5-FU )＞ Mitoxantrone ( MIT )＞Vincristine ( VCR )＞ Pirarubicin ( THP )＞ Isosophosphamide ( IFO )＞ Carboplatin ( CBP )＞Cyclophosphamide ( CTX)＞ Methotrexate ( MTX)＞ Changchun Rui bin ( NVB) . The sensitivity of chemotherapy regimens in the three groups from high to low was docetaxel/doxorubicin/cyclophosphamide (TAC )＞cyclophosphamide/epirubicin/fluorouracil ( CEF )＞cyclophosphamide/methotrexate/fluorouracil (CMF). The sensitivity rates of PTX,EPI and DDP were 54%(104/192),42%(81/192) and 37%(71/192) respectively. (2) The average inhibitory rates of DDP,CBP and MIT in stage III breast cancer was higher than those in stage I and II breast cancer,and the differences were statistically significant ( F＝11. 14,4. 303,3. 182,P＜0. 05). (3) HR-breast cancer is more sensitive than HR+breast cancer,PTX, EPI,THP,MIT in HER-2(+) breast cancer is more sensitive than in HER-2(－) breast cancer. Conclusion As a widely used drug sensitivity test method, MTT assay has a certain reference value for screening sensitive drugs and selecting clinical chemotherapy regimens in neoadjuvant chemotherapy of breast cancer. PTX,EPI and DDP are more sensitive to other breast cancer cells than other drugs. Chemotherapy based on in vitro susceptibility results improves the efficiency of chemotherapy and decreases the proportion of changes in chemotherapy schemes due to inefficiency.

Clinical outcome of therapeutic cervical cerclage in short cervix syndrome / 中华妇产科杂志

Objective To investigate the clinical effect of therapeutic cervical cerclage on short cervix syndrome for anti-premature birth in the second trimester. Methods Totally 44 singleton pregnant patients were diagnosed as short cervix syndrome, which was cervical length ≤2.5 cm without cervical dilatation,and received treatment from January 2008 and July 2015 in Peking University Third Hospital were collected. Among them, 30 patients who received therapeutic cervical cerclage were defined as cerclage group and another 14 cases who received conservative treatment were defined as un-cerclage group. The days of conservative treatment, delivery rate of different gestational weeks, birth weight of newborns, neonatal survival rate within 7 days of birth were analyzed between the two groups. Results There were no significant differences between the two groups in days of pregnancy conservative treatment [103(84-141)vs 105(85-114)days], delivery weeks [38.0(35.5-39.4)vs 38.5(37.3-39.5)weeks], birth weight of newborns [3120(2750-3400)vs 3130(2760-3545)g], and survival rate of newborns [100％(30/30)vs 13/14]. The fetuses of both groups were all delivered after 28 weeks. There was no significant difference in accumulated delivery rate between the two groups after 32 weeks, 34 weeks, and 37 weeks, respectively(all P>0.05). Conclusions The treatment of cervical cerclage is not superior to conservative means in single pregnancy of cervical length ≤2.5 cm without cervical dilatation. For such patients with short cervix syndrome, the treatment of cervical cerclage may not be necessary, but dynamic monitoring and search for the causing factors and prompt treatment are more important.

Clinical efficacy of implementation of standardized enteral nutrition in patients with severe traumatic brain injury undergoing mechanical ventilation / 中国中西医结合急救杂志

Objective To investigate the clinical efficacy of implementation of standardized enteral nutrition (EN) and its effects on prognosis in patients with severe traumatic brain injury (sTBI) undergoing mechanical ventilation (MV). Methods Eighty-eight patients with sTBI undergoing MV admitted to the Department of Critical Care Medicine of Yuyao People's Hospital from January 2016 to December 2017 were enrolled, they were divided into a control group (42 cases) and an experiment group (46 cases) depending on the demarcation timing of January 1, 2017, the beginning time of implementing standardized EN. All the patients received early EN and conventional treatment in the two groups. Additionally, the procedure of standardized EN was implemented in the experiment group. The differences in starting time of EN, the first defecation time, the rates of EN therapeutic energy and protein supply reaching their respective targets, duration of MV and ICU stay and 28-day mortality were compared between the two groups. Results The starting time of EN (hours: 25.61±8.74 vs. 32.79±8.63) and first defecation time (days: 3.03±0.79 vs. 3.61±0.89) were significantly earlier in the experiment group than those in the control group (both P < 0.05); the rates of energy and protein supply reaching the respective targets on the 5th day and 7th day after receiving EN were all significantly higher in the experiment group than those in the control group [rates of energy supply reaching target on the 5th day: (44.83±13.99)% vs. 37.59±10.88, and on the 7th day: (68.07±10.68)% vs. (62.69±9.87)%; rate of protein supply reaching target on the 5th day: (31.93±9.49)% vs. (27.06±8.08)%, and on the 7th day: (62.09±9.91)% vs. (54.55±11.27) %, all P < 0.05]; the durations of MV (hours: 9.24±2.91 vs. 10.67±3.41) and ICU stay (days: 12.09±3.37 vs. 13.93±4.98) in the experiment group were significantly shorter than those in the control group (all P < 0.05). No statistical significant difference in the 28-day mortality was observed between the experiment group and control group [21.74% (10/46) vs. 19.05% (8/42), P > 0.05]. Conclusion The efficacy of implementation of standardized EN in patients with sTBI undergoing MV is very significant, as it can significantly improve the rate of reaching EN target, and shorten the duration of MV and ICU stay.

Research progress of jasmonate-responsive transcription factors in regulating plant secondary metabolism / 中国中药杂志

Jasmonates, as a plant endogenous hormone, can induce the biosynthesis of terpenoids, alkaloids and flavonoids and other medicinal active ingredients, and play an important role in the plant secondary metabolic process. The tanscription factors can activate the expression of multiple genes in the biosynthesis of plant secondary metabolites by binding the cis-elements of the target genes. Then, it effectively activates or inhibits the activities of the enzymes on the biosynthesis of plant secondary metabolites, further regulate specific biosynthesis and accumulation of secondary metabolites. Here, We review recent major progresses regarding the regulation of secondary metabolites by JAs-responsive transcription factors (TFs) (including AP2/ERFs, bHLH, MYB and WRKY). That provides suggestions for further analysis of jasmonic acid signaling pathway and regulation of secondary metabolism, and explores the potential value of transcription factor in improving the medicinal active ingredients.

Construction and eukaryotic expression of recombinant HSA-TP5 fusion gene expression vector / 吉林大学学报(医学版)

Objective:To construct the expression vector of the fusion protein of human serum albumin (HSA) and thymopentin (TP5) and to express it in Pichia pastoris,and to elucidate the biological activity of fusion protein.Methods:The HSA-TP5 fusion gene was constructed by gene recombination and transfected into Pichia pastoris to construct the eukaryotic expression system of HSA-TP5.The recombinant eukaryotic expression plasmid of PPICZα-HSA-TP5 was obtained by agarose gel electrophoresis and purification reagent.The two step fermentation method was used to ferment gene engineering bacteria of HSA-TP5 in high density,and the fermentation supernatant protein was precipitated and concentrated;the purified fusion protein was obtained by cation exchange chromatography and hydrophobic chromatography and analyzed by SDS-polyacrylamide gel electrophoresis.The effect of the fusion protein on the proliferation of lymphocytes was detected by MTT assay.Results:The HSA target gene fragment with length of 1 845 bp was achieved by PCR method.The HSA-TP5-pPICZαC fusion plasmid was identified by restriction endonuclease digestion,and the fragment length was 707 bp.The sequence analysis showed that the HSA and TP5 sequences of the target genes were identical with the gene sequences reported in GenBank and were fused by forward fusion.PCR method confirmed that the eukaryotic recombinant plasmid PPICZ αC-HSA-TP5 was integrated into the yeast genome,and compared with control group,the target gene PCR product length was found to be 1 860 bp.SDS-PAGE analysis showed that the expression level of HSA-TP5 fusion protein was gradually increased with the induction time within 72 h.HSA-TP5 fusion protein was purified by cation exchange chromatography and AKTA multifunctional protein purification system.The MTT assay results showed that HSA-TP5 fusion protein was consistent with TP5 protein in promoting lymphocyte proliferation activity.Conclusion:HSA-TP5 fusion protein can be obtained by constructing the eukaryotic expression system of Pichia pastoris and owns the biological activity.

Effect of blood pressure control level on perinatal outcomes in women with mild-moderate gestational hypertension / 中华妇产科杂志

Objective To investigate the effect of blood pressure (BP) control level on perinatal outcomes in women with mild-moderate gestational hypertension (GHp). Methods Totally,344 women diagnosed initially as mild-moderate GHp who delivered in Peking University Third Hospital from January 2012 to December 2016 were recruited. They were divided into four groups according to the stabilized level of BP during pregnancy. (1) Group A:BP<130/80 mmHg(1 mmHg=0.133 kPa);(2) Group B:BP(130-139)/(80-89) mmHg; (3) Group C: BP (140-149)/(90-99) mmHg; (4) Group D: BP (150-159)/(100-109)mmHg. The clinical profile and incidence of severe GHp, pre-eclampsia with proteinuria (PE+Upro), severe pre-eclampsia (sPE), small-for-gestational age (SGA) were compared among the four groups. Student t-test was preformed to normal distributive data and Kruskal-Wallis test was used to non-normally distributed variables. Chi-square test was used in count data. Logistic regression analysis was adopted for multiple-factor analysis. Results (1) The incidence of severe GHp in group A was lower than group B (P<0.05). The incidences of severe GHp and sPE in the group B was lower than those in group C (P<0.05). While there was no difference in the incidence of PE+Upro and SGA among the four groups (P>0.05). And the incidence of severe GHp in group D had no difference with group A, B, C (P>0.05). (2) In the 48 patients who used medications to control BP, the occurence of severe GHp in those whose initial BP was(140-149)/(90-99)mmHg was lower than those of≥160/110 mmHg (P<0.05). But the incidence of severe GHp had no significant difference between patients whose initial BP was(140-149)/(90-99)mmHg and patients whose initial BP was(150-159)/(100-109)mmHg (P>0.05). The initial BP level had no impact on the incidence of PE+Upro, sPE and SGA (P>0.05). (3) Multivariate logistic regression analysis showed that the BP level before using medications (OR=3.566, 95%CI:1.080-11.771, P=0.037) and the BP level maintained (OR=4.787, 95%CI:1.115-20.551,P=0.035) were independent factor that affected the incidence of severe GHp. Edema (OR=2.651, 95%CI:1.628-4.316 P=0.000), fetal growth restriction(FGR;OR=1.103, 95%CI:1.427-5.914,P=0.002)and the onset gestational age of GHp (OR=0.755, 95%CI:0.578-0.985,P=0.038) were independent factors that affected the incidence of PE+Upro. The tendency of FGR (OR=17.787, 95%CI:1.833-40.396 P=0.000), history of PE (OR=5.294, 95%CI:1.086-25.800,P=0.039) and the BP level during pregnancy (OR=2.109, 95%CI:1.274-3.491,P=0.004) were independent factors affecting the incidence of sPE. FGR tendency was independent factor affecting the incidence of SGA (OR=25.622, 95%CI:2.596-252.864,P=0.005). Conclusion A satisfied control of BP is helpful to reduce severe GHp and sPE, but the incidence of SGA does not affected.

Construction and eukaryotic expression of recombinant HSA-TP5 fusion gene expression vector / 吉林大学学报(医学版)

Objective:To construct the expression vector of the fusion protein of human serum albumin (HSA) and thymopentin (TP5) and to express it in Pichia pastoris,and to elucidate the biological activity of fusion protein.Methods:The HSA-TP5 fusion gene was constructed by gene recombination and transfected into Pichia pastoris to construct the eukaryotic expression system of HSA-TP5.The recombinant eukaryotic expression plasmid of PPICZα-HSA-TP5 was obtained by agarose gel electrophoresis and purification reagent.The two step fermentation method was used to ferment gene engineering bacteria of HSA-TP5 in high density,and the fermentation supernatant protein was precipitated and concentrated;the purified fusion protein was obtained by cation exchange chromatography and hydrophobic chromatography and analyzed by SDS-polyacrylamide gel electrophoresis.The effect of the fusion protein on the proliferation of lymphocytes was detected by MTT assay.Results:The HSA target gene fragment with length of 1 845 bp was achieved by PCR method.The HSA-TP5-pPICZαC fusion plasmid was identified by restriction endonuclease digestion,and the fragment length was 707 bp.The sequence analysis showed that the HSA and TP5 sequences of the target genes were identical with the gene sequences reported in GenBank and were fused by forward fusion.PCR method confirmed that the eukaryotic recombinant plasmid PPICZ αC-HSA-TP5 was integrated into the yeast genome,and compared with control group,the target gene PCR product length was found to be 1 860 bp.SDS-PAGE analysis showed that the expression level of HSA-TP5 fusion protein was gradually increased with the induction time within 72 h.HSA-TP5 fusion protein was purified by cation exchange chromatography and AKTA multifunctional protein purification system.The MTT assay results showed that HSA-TP5 fusion protein was consistent with TP5 protein in promoting lymphocyte proliferation activity.Conclusion:HSA-TP5 fusion protein can be obtained by constructing the eukaryotic expression system of Pichia pastoris and owns the biological activity.

Effect of blood pressure control level on perinatal outcomes in women with mild-moderate gestational hypertension / 中华妇产科杂志

Objective To investigate the effect of blood pressure (BP) control level on perinatal outcomes in women with mild-moderate gestational hypertension (GHp). Methods Totally,344 women diagnosed initially as mild-moderate GHp who delivered in Peking University Third Hospital from January 2012 to December 2016 were recruited. They were divided into four groups according to the stabilized level of BP during pregnancy. (1) Group A:BP<130/80 mmHg(1 mmHg=0.133 kPa);(2) Group B:BP(130-139)/(80-89) mmHg; (3) Group C: BP (140-149)/(90-99) mmHg; (4) Group D: BP (150-159)/(100-109)mmHg. The clinical profile and incidence of severe GHp, pre-eclampsia with proteinuria (PE+Upro), severe pre-eclampsia (sPE), small-for-gestational age (SGA) were compared among the four groups. Student t-test was preformed to normal distributive data and Kruskal-Wallis test was used to non-normally distributed variables. Chi-square test was used in count data. Logistic regression analysis was adopted for multiple-factor analysis. Results (1) The incidence of severe GHp in group A was lower than group B (P<0.05). The incidences of severe GHp and sPE in the group B was lower than those in group C (P<0.05). While there was no difference in the incidence of PE+Upro and SGA among the four groups (P>0.05). And the incidence of severe GHp in group D had no difference with group A, B, C (P>0.05). (2) In the 48 patients who used medications to control BP, the occurence of severe GHp in those whose initial BP was(140-149)/(90-99)mmHg was lower than those of≥160/110 mmHg (P<0.05). But the incidence of severe GHp had no significant difference between patients whose initial BP was(140-149)/(90-99)mmHg and patients whose initial BP was(150-159)/(100-109)mmHg (P>0.05). The initial BP level had no impact on the incidence of PE+Upro, sPE and SGA (P>0.05). (3) Multivariate logistic regression analysis showed that the BP level before using medications (OR=3.566, 95%CI:1.080-11.771, P=0.037) and the BP level maintained (OR=4.787, 95%CI:1.115-20.551,P=0.035) were independent factor that affected the incidence of severe GHp. Edema (OR=2.651, 95%CI:1.628-4.316 P=0.000), fetal growth restriction(FGR;OR=1.103, 95%CI:1.427-5.914,P=0.002)and the onset gestational age of GHp (OR=0.755, 95%CI:0.578-0.985,P=0.038) were independent factors that affected the incidence of PE+Upro. The tendency of FGR (OR=17.787, 95%CI:1.833-40.396 P=0.000), history of PE (OR=5.294, 95%CI:1.086-25.800,P=0.039) and the BP level during pregnancy (OR=2.109, 95%CI:1.274-3.491,P=0.004) were independent factors affecting the incidence of sPE. FGR tendency was independent factor affecting the incidence of SGA (OR=25.622, 95%CI:2.596-252.864,P=0.005). Conclusion A satisfied control of BP is helpful to reduce severe GHp and sPE, but the incidence of SGA does not affected.

Bioinformatics analysis of geranylgeranyl pyrophosphate synthase coding gene and amino acid sequence in Lamiaceae / 中国中药杂志

Geranylgeranyl pyrophosphate synthase enzyme is one of the key enzymes in the synthesis pathway of diterpenoid. Nine Lamiaceae genus GGPS synthase in Genebank was analyzed in this article. GGPS synthase the nucleic acid sequences and amino acid sequences, physicochemical properties, the signal peptide, leader peptides, transmembrane topological structure, hydrophobic, hydrophilic, subcellular localization, secondary structure, function domain, tertiary structure and evolutional relationship were predicted by using bioinformatics methods.Phylogenetic tree was constructed for the geranylgeranyl pyrophosphate synthase enzyme protein family. The results showed that GGPS amino acid sequence of the physical and chemical properties were basically identical, mainly hydrophilic protein, there existed chloroplast transit peptide, and no signal peptide and membrane structure domain, which mainly located in the chloroplast, the minor part located in mitochondria. The main secondary structures of the proteins are alpha helix and random coil. All these proteins have catalytic residues, aspartate-rich region, active site lid residues, substrate-Mg2+ binding site. The results provide theoretical reference for study on both the enzymatic characteristics of GGPS and the biosynthesis pathway of diterpenoid.

Cloning, subcellular localization, and heterologous expression of ApNAC1 gene from Andrographis paniculata / 中国中药杂志

Andrographis paniculata is widely used as medicinal herb in China for a long time and andrographolide is its main medicinal constituent. To investigate the underlying andrographolide biosynthesis mechanisms, RNA-seq for A. paniculata leaves with MeJA treatment was performed. In A. paniculata transcriptomic data, the expression pattern of one member of NAC transcription factor family (ApNAC1) matched with andrographolide accumulation. The coding sequence of ApNAC1 was cloned by RT-PCR, and GenBank accession number was KY196416. The analysis of bioinformatics showed that the gene encodes a peptide of 323 amino acids, with a predicted relative molecular weight of 35.9 kDa and isoelectric point of 6.14. To confirm the subcellular localization, ApNAC1-GFP was transiently expressed in A. paniculata protoplast. The results indicated that ApNAC1 is a nucleus-localized protein. The analysis of real-time quantitative PCR revealed that ApNAC1 gene predominantly expresses in leaves. Compared with control sample, its expression abundance sharply increased with methyl jasmonate treatment. Based on its expression pattern, ApNAC1 gene might involve in andrographolide biosynthesis. ApNAC1 was heterologously expressed in Escherichia coli and recombinant protein was purified by Ni-NTA agarose. Further study will help us to understand the function of ApNAC1 in andrographolide biosynthesis.