ObjectiveThrough qualitatively and quantitatively analysis of the differences in chemical composition between the combined decoction and single decoction of Huaganjian and comparison of their core efficacy， to explore the rationality of the flexible clinical application of Huaganjian compound preparations and single-flavored dispensing granules. MethodsUltra performance liquid chromatography-quadrupole-electrostatic field orbitrap high resolution mass spectrometry（UPLC-Q-Exactive Orbitrap MS） was used to qualitatively analyze the combined decoction and single decoction samples of Huaganjian， and meanwhile， the contents of four index components（geniposide， paeoniflorin， hesperidin and paeonol） were quantitatively analyzed by high performance liquid chromatography（HPLC）. Nonalcoholic fatty liver disease（NAFLD） rat model induced by high-fat diet was applied to compare the efficacy of combined decoction and single decoction of Huaganjian. A total of 30 male SD rats were randomly divided into the control group， model group， lovastatin group（1.8 mg·kg^-1）， combined decoction group（1.26 g·kg^-1） and single decoction group（1.18 g·kg^-1）. After successful modeling， lovastatin group， combined decoction group and single decoction group were given corresponding doses of drugs by intragastric administration every day， and the control group and model group were given equal amounts of normal saline by intragastric administration， after 4 weeks of administration， the serum and liver tissues were collected， and the contents of alanine aminotransferase（ALT）， aspartate aminotransferase（AST）， total cholesterol（TC）， triglyceride（TG）， low-density lipoprotein cholesterol（LDL-C） and high-density lipoprotein cholesterol（HDL-C） in serum of rats were detected， and the liver pathological examination was carried out by hematoxylin-eosin（HE） staining and oil red O staining， so as to compare differences of their efficacy. ResultsSeventy chemical components were initially identified and attributed from the lyophilized powder of the combined decoction and single decoction samples of Huaganjian， and there was no obvious difference in composition between the two. Further quantitative analysis showed that the contents of geniposide， paeoniflorin， hesperidin and paeonol in the combined decoction samples were significantly increased when compared with those of the single decoction samples（P<0.01）. The pharmacodynamic results showed that compared with the model group， both the combined and single decoction groups of Huaganjian could improve the liver index of NAFLD rats， reduce the serum levels of AST， ALT， TC， TG and LDL-C， increase the serum level of HDL-C， and ameliorate the pathological changes of liver cell steatosis and fat accumulation. However， there was no significant difference in pharmacodynamic effects between the combined decoction group and the single decoction group. ConclusionThere is no significant difference between the combined decoction and single decoction of Huaganjian in terms of chemical composition， but the contents of the four index components show significantly difference. Both of them can significantly improve the fat accumulation and liver function in NAFLD rats. This study provides a reference basis for the rational clinical application and evaluation of famous classical formula compound preparations and single-flavored dispensing granules.

Objective To investigate the role and regulatory mechanism of stress-inducing protein 1(SESN1)in liver gluconeogenesis of fasting mice.Methods RT-qPCR was used to detect mRNA expression of SESN1 in liver tissues of C57BL/6J mice and primary mouse hepatocytes treated with forskolin(Fsk)and dexamethasone(Dex).HepG2 cells were transfected with plasmids and the effects of SESN1 overexpression on mRNA expression of gluconeogenesis related genes PGC-1α,PEPCK and G6Pase was detected by RT-qPCR.The effect of SESN1 on the promoter activity of PGC-1α in HepG2 cells was studied using a dual luciferase reporter system.The effect of SESN1 on PGC-1α deacetylation was detected by overexpression of SESN1 and inhibition of SIRT1 expression.By knocking down SIRT1 expression,we detected whether it mediated the changes in mRNA levels of SESN1 in-duced gluconeogenesis related genes.Results The mRNA expression of SESN1 was significantly increased in liver tissues of starved C57BL/6J mice and in primary hepatocytes treated with Fsk and Dex(P<0.001).Over-expression of SESN1 in HepG2 cells promoted mRNA expression of PGC-1α,PEPCK and G6Pase(P<0.001)and promoter activity of PGC-1α(P<0.001).Over-expression of SESN1 decreased the acetylation level of PGC-1α in primary hepatocytes.Sirt family inhibitors NAM and shRNA adenovirus interfered with SIRT1 expression respective-ly,and antagonized the deacetylation effect of SESN1 on PGC-1α.The expression of PGC-1α,PEPCK and G6Pase induced by SIRT1 was also significantly impaired(P<0.000 1).Conclusions SESN1 regulates liver gluconeogene-sis in mice with a SIRT1-dependent mechanism.

Objective To investigate the relationship between left ventricular hypertrophy(LVH)diagnosed by Peguero-Lo-Presti index and recurrence of paroxysmal atrial fibrillation(AF)after radiofrequency ablation.Methods A total of 652 patients with paroxysmal atrial fibrillation who underwent radiofrequency ablation were selected.According to Peguero-Lo-Presti index,patients were divided into the LVH group(167 cases)and the normal left ventricle group(485 cases).Baseline data were collected,and regular follow-up was performed at 3,6 and 12 months after radiofrequency catheter ablation.The recurrence of AF was assessed.Kaplan-Meier survival curve was used to analyze the recurrence rate of AF in the two groups.Cox proportional hazard model was used to assess risk factors for recurrent atrial fibrillation.Results The median follow-up time was 20.5(15.0,26.0)months.A total of 155 patients(23.8%)developed recurrence of AF,including 95 patients in the LVH group and 60 patients in the LVN group.The recurrence rate without AF was significantly lower in the LVH group than that in the LVN group(64.1%vs.80.4%,Log-rank χ2=26.361,P＜0.01).After adjusting for age,sex,body mass index,hypertension,diabetes,coronary heart disease,cardiac dysfunction,left anteroposterior and posterior atrial diameter,left ventricular end-diastolic diameter,and left ventricular ejection fraction,LVH diagnosed by Peguero-Lo-Presti index was still a risk factor for recurrent AF[HR(95%CI):2.359(1.663-3.345),P＜0.01].Conclusion In patients with paroxysmal AF,LVH diagnosed by Peguero-Lo-Presti index is a risk factor of AF recurrence after radiofrequency catheter ablation.

Objective:To investigate the alteration of right ventricular function after catheter ablation in patients with persistent atrial fibrillation(PAF) and concomitant heart failure with preserved ejection fraction (HFpEF).Methods:The prospective observational study was performed in patients with HFpEF-PAF and undergoing first-time radiofrequency ablation procedures in the First Affiliated Hospital of Nanjing University between May to December 2019. Right ventricular functional parameters were measured before and 5 days, 1, 3, 6 and 12 months after the ablation by transthoracic echocardiography, respectively, including the right ventricular fractional area change (RVFAC), tricuspid annular plane systolic excursion(TAPSE), tricuspid annular diameter (TVAD), tricuspid annular peak systolic speed(TDI-S′) and longitudinal strain of right ventricular free wall (RVFLS). Meanwhile, routine ECG and Holter recordings were performed at each follow-up time point.Results:In this study, atrial fibrillation (AF) recurrence occurred in 4 patients at the 3rd month after ablation, and 7 patients failed to follow up due to the Covid-19. Finally, 19 patients were followed up for the evaluation of cardiac function after catheter ablation. Compared with pre-ablation, right ventricular structural and functional paramters(RVFAC, TAPSE, TVAD, TDI-S′, RVFLS) improved significantly at all stages of follow-up( all P<0.05). Patients with atrial fibrillation recurrence had lower RVFLS and TDI-S′ at the baseline( P=0.039, P=0.019). Conclusions:Right ventricular function could improve in HFpEF-PAF patients who maintain sinus rhythm after radiofrequency ablation.

Proteins and peptides have become a significant therapeutic modality for various diseases because of their high potency and specificity. However, the inherent properties of these drugs, such as large molecular weight, poor stability, and conformational flexibility, make them difficult to be formulated and delivered. Injection is the primary route for clinical administration of protein and peptide drugs, which usually leads to poor patient's compliance. As a portable, minimally invasive device, microneedles (MNs) can overcome the skin barrier and generate reversible microchannels for effective macromolecule permeation. In this review, we highlighted the recent advances in MNs-mediated transdermal delivery of protein and peptide drugs. Emphasis was given to the latest development in representative MNs design and fabrication. We also summarize the current application status of MNs-mediated transdermal protein and peptide delivery, especially in the field of infectious disease, diabetes, cancer, and other disease therapy. Finally, the current status of clinical translation and a perspective on future development are also provided.

Leucine dehydrogenase (LDH) is the key rate-limiting enzyme in the production of L-2-aminobutyric acid (L-2-ABA). In this study, we modified the C-terminal Loop region of this enzyme to improve the specific enzyme activity and stability for efficient synthesis of L-2-ABA. Using molecular dynamics simulation of LDH, we analyzed the change of root mean square fluctuation (RMSF), rationally designed the Loop region with greatly fluctuated RMSF, and obtained a mutant EsLDHD2 with a specific enzyme activity 23.2% higher than that of the wild type. Since the rate of the threonine deaminase-catalyzed reaction converting L-threonine into 2-ketobutyrate was so fast, the multi-enzyme cascade catalysis system became unbalanced. Therefore, the LDH and the formate dehydrogenase were double copied in a new construct E. coli BL21/pACYCDuet-RM. Compared with E. coli BL21/pACYCDuet-RO, the molar conversion rate of L-2-ABA increased by 74.6%. The whole cell biotransformation conditions were optimized and the optimal pH, temperature and substrate concentration were 7.5, 35 °C and 80 g/L, respectively. Under these conditions, the molar conversion rate was higher than 99%. Finally, 80 g and 40 g L-threonine were consecutively fed into a 1 L reaction mixture under the optimal conversion conditions, producing 97.9 g L-2-ABA. Thus, this strategy provides a green and efficient synthesis of L-2-ABA, and has great industrial application potential.
Aminobutyrates ; Escherichia coli/genetics* ; Leucine Dehydrogenase/genetics* ; Threonine Dehydratase

Objective To analyze the residents' iodine nutrition level of Yunfu City in Guangdong Province from 2009 to 2016,in order to provide evidence for making prevention strategy of iodine deficiency disorders.Methods Four surveys on salt iodine levels,four surveys on urinary iodine levels of children aged 8-10 years old,two surveys on urinary iodine levels of pregnant women and one survey on iodine content in drinking water were carried out according to the prevention and control planning for endemic disease in Guangdong Province in Yunfu City from 2009 to 2016.Salt iodine was determined by direct titration (GB/T 13025.7-1999,GB/T 13025.7-2012),urinary iodine was determined by arsenic cerium catalytic spectrophotometry (WS/T 107-2006,WS/T 107.1-2016),water iodine was determined by the national iodine deficiency disease reference laboratory recommended method.Results In 2009,2011,2015 and 2016,1 464,1 464,1 500,1 500 salt samples and 500,388,1 000,1 000 children were investigated,the medians of salt iodine levels were 30.3,28.1,24.8,and 24.8 mg/kg,respectively,and the medians of children urinary iodine levels were 208.3,188.3,143.1 and 165.2 μg/L,respectively.Five hundred,five hundred pregnant women were investigated in 2015 and 2016,and the medians of pregnant women urinary iodine levels were 101.0 and 96.4 μg/L,respectively.A total of 1 149 samples of drinking water were investigated in 2011-2016,the median of water iodine was 2.8 μg/L.Conclusion Yunfu is an iodine deficiency city,after adjusting the iodine content of edible salt in 2012,iodine nutrition in children is at an appropriate level,but the iodine intake level of pregnant women is relatively lower,so prevention and treatment of iodine deficiency disorders and urinary iodine monitoring should be strengthened in pregnant women.

Objective To investigate influence of different doses of dexmedetomidine on hemodynamics of pregnancy-induced hypertension syndrome (PIH) patients with cesarean section.Methods A total of 325 PIH cases in our hospital were divided into control group and A,B,C,D groups,with 65 cases in each group.The control group received only basic anesthesia,A,B,C,D groups were given dexmedetomidine intravenous infusion of 0.1,0.2,0.3,0.4 μg/(kg · h),respectively.Systolic blood pressure (SBP),diastolic blood pressure (DBP),heart rate (HR) and other indicators before epidural administration (T0),skin incision (T1),fetal delivery (T2),at the end of surgery (T3) were recorded.Results SBP,DBP,HR of 4 time points in 5 groups had a significant difference (P ＜ 0.05).At T0,SBP,DBP and HR of 5 groups were significantly higher than that of T1,T2,T3 (P ＜ 0.05).At T1,SBP,DBP,HR were significantly higher than that of T2 and T3 (P ＜ 0.05),At T2,SBP,DBP,HR of 5 groups were significantly higher than T3 (P ＜ 0.05).At T0,SBP,DBP,HR among 5 groups showed no significant difference (P ＞ 0.05).At T1 ～ T3,DBP,HR SBP of the 5 groups had a significant difference (P ＜ 0.05),SBP DBP and HR in D group were significantly lower than that of A,B,C groups,and control group (P ＜ 0.05).SBP,DBP,HR in C group were significantly lower than B group,and the control group (P ＜ 0.05).SBP,DBP,HR in B group were significantly lower than A group and the control group (P ＜ 0.0 5);SBP,DBP,HR between A group and control group showed no significant difference (P ＞ 0.05).Conclusion The dose of 0.4 μg/(kg · h) for PIH patients with cesarean section has stable hemodynamics.With the increase of dexmedetomidine dose,the effect is more obvious.

Objective To investigate influence of different doses of dexmedetomidine on hemodynamics of pregnancy-induced hypertension syndrome (PIH) patients with cesarean section.Methods A total of 325 PIH cases in our hospital were divided into control group and A,B,C,D groups,with 65 cases in each group.The control group received only basic anesthesia,A,B,C,D groups were given dexmedetomidine intravenous infusion of 0.1,0.2,0.3,0.4 μg/(kg · h),respectively.Systolic blood pressure (SBP),diastolic blood pressure (DBP),heart rate (HR) and other indicators before epidural administration (T0),skin incision (T1),fetal delivery (T2),at the end of surgery (T3) were recorded.Results SBP,DBP,HR of 4 time points in 5 groups had a significant difference (P ＜ 0.05).At T0,SBP,DBP and HR of 5 groups were significantly higher than that of T1,T2,T3 (P ＜ 0.05).At T1,SBP,DBP,HR were significantly higher than that of T2 and T3 (P ＜ 0.05),At T2,SBP,DBP,HR of 5 groups were significantly higher than T3 (P ＜ 0.05).At T0,SBP,DBP,HR among 5 groups showed no significant difference (P ＞ 0.05).At T1 ～ T3,DBP,HR SBP of the 5 groups had a significant difference (P ＜ 0.05),SBP DBP and HR in D group were significantly lower than that of A,B,C groups,and control group (P ＜ 0.05).SBP,DBP,HR in C group were significantly lower than B group,and the control group (P ＜ 0.05).SBP,DBP,HR in B group were significantly lower than A group and the control group (P ＜ 0.0 5);SBP,DBP,HR between A group and control group showed no significant difference (P ＞ 0.05).Conclusion The dose of 0.4 μg/(kg · h) for PIH patients with cesarean section has stable hemodynamics.With the increase of dexmedetomidine dose,the effect is more obvious.

We constructed plasmid pMTac to overexpress 3-ketosteroid-Δ1-dehydrogenase (KSDD) in Mycobacterium neoaurum JC-12 for improving androst-1,4-diene-3,17-dione (ADD) production. To construct pMTac, pACE promoter on pMF41 was replaced by tac promoter, and then four recombinants were constructed, which were M. neoaurum JC-12/pMF41-gfp, M. neoaurum JC-12/pMTac-gfp, M. neoaurum JC-12/pMF41-ksdd and M. neoaurum JC-12/pMTac-ksdd. Fluorescence detection results show that much more green fluorescent protein (GFP) was expressed in M. neoaurum JC-12/pMTac-ksdd than M. neoaurum JC-12/pMF41-ksdd. The activity of KSDD was 2.41 U/mg in M. neoaurum JC-12/pMTac-ksdd, 6.53-fold as that of M. neoaurum JC-12 and 4.36-fold as that of M. neoaurum JC-12/pMF41-ksdd. In shake flask fermentation, ADD production of M. neoaurum JC-12/pMTac-ksdd was 5.94 g/L, increased about 22.2% compared to the original strain M. neoaurum JC-12 and 12.7% to M. neoaurum JC-12/pMF41-ksdd. AD (4-androstene-3,17-dione) production of JC-12/pMTac-ksdd was 0.17 g/L, decreased 81.5% compared to M. neoaurum JC-12 and 71.2% to M neoaurum JC-12/pMF41-ksdd. In the 5 L fermenter, 20 g/L phytosterols was used as substrate, ADD production of M. neoaurum JC-12/pMTac-ksdd was improved to 10.28 g/L. pMTac is favorable for expressing KSDD in M. neoaurum JC-12, and overexpression of KSDD has beneficial effect on ADD producing, and it is the highest level ever reported using fermentation method in M. neoaurum.
Androstadienes ; metabolism ; Fermentation ; Industrial Microbiology ; Mycobacterium ; Oxidoreductases ; genetics ; metabolism ; Phytosterols ; metabolism ; Plasmids