ObjectiveThe U6 promoter is an essential element for driving sgRNA expression in the clustered regularly interspaced short palindromic repeat sequences/CRISPR-associated protein 9（CRISPR/Cas9）gene editing system in dicotyledonous plants. Endogenous U6 promoters typically exhibit higher transcriptional activity， which can significantly improve gene editing efficiency. This study aims to identify endogenous U6 promoters in Artemisia annua to optimize its CRISPR/Cas9 gene editing system， which holds significant importance for its molecular breeding. MethodsOn the basis of the highly conserved U6 snRNA sequences in Arabidopsis thaliana， endogenous U6 promoters were screened in the A. annua genome. Expression vectors were constructed with candidate AaU6 promoter driving the firefly luciferase （LUC） reporter gene， and then transiently transformed into Nicotiana benthamiana. Transcriptional activities of the promoters were measured and compared by in vivo imaging and the Dual Luciferase Reporter assay. ResultsEight endogenous U6 promoters were successfully cloned from A. annua. Sequences alignment revealed that all these promoters contained the two conserved cis-acting elements， upstream sequence element （USE） and TATA-box， which affected their transcriptional activity. Dual-luciferase activity assays indicated that the transcriptional activities of AaU6-3， AaU6-1， and AaU6-5 were significantly higher than that of the Arabidopsis AtU6-26 promoter， with AaU6-3 exhibiting the highest activity. ConclusionThis study identified three endogenous AaU6 promoters with high transcriptional activity in A. annua， providing key functional elements for establishing an efficient gene editing system in A. annua. These findings will contribute to advancing precision molecular breeding and high-quality germplasm innovation in A. annua.

Antibiotic resistance is spreading at a faster rate than new antibiotic agents applied for clinical remedies. It is an urgent need to discover potential compounds to combat multidrug-resistant (MDR) bacteria. Marine fungi offer a promising avenue for mining antibiotic-like molecules with chemical diversity. To discover structurally novel and antibiotic metabolites, we screened the in-house marine fungus genome library and found a fungus Stephanonectria keithii LZD-10-1 containing a non-ribosomal peptide synthetase (NRPS) cluster with 18 modules to synthesize a new subfamily of peptaibols with effective eradication against MDR pathogens. Targeting isolation of the cultured fungus afforded six new peptaibols, which exhibit the ability to kill MDR bacteria by targeting bacterial membrane phospholipids, especially phosphatidylglycerol (PG), leading to the dysfunction of bacterial membranes. Furthermore, their efficacies against methicillin-resistant Staphylococcus aureus (MRSA) in both Galleria mellonella and mouse wound infection models were observed. This study underscores the significance of employing genome-guided approaches to identify untapped marine fungi as potential sources for novel antibiotic candidates with unique scaffolds.

Grooming,as an evolutionarily conserved repetitive behavior,is common in various animals,including humans,and serves essential functions including,but not limited to,hygiene maintenance,thermoregulation,de-arousal,stress reduction,and social behaviors.In rodents,grooming involves a patterned and sequenced structure,known as the syntactic chain with four phases that comprise repeated stereotyped movements happening in a cephalocaudal progression style,beginning from the nose to the face,to the head,and finally ending with body licking.The context-dependent occurrence of grooming behavior indicates its adaptive significance.This review briefly summarizes the neural substrates responsible for rodent grooming behavior and explores its relevance in rodent models of neuropsychiatric disorders and neurodegenerative diseases with aberrant grooming phenotypes.We further emphasize the utility of rodent grooming as a reliable measure of repetitive behavior in neuropsychiatric models,holding promise for translational psychiatry.Herein,we mainly focus on rodent self-grooming.Allogrooming(grooming being applied on one animal by its conspecifics via licking or carefully nibbling)and heterogrooming(a form of grooming behavior directing towards another animal,which occurs in other contexts,such as maternal,sexual,aggressive,or social behaviors)are not covered due to space constraints.

Objective: To evaluate the effects of Liu Zi Jue Qigong (LQG)-based breathing training on the curvature and range of motion (ROM) of the thoracic and lumbar spines in different positions. Methods: Forty-seven college students were selected as subjects and randomly divided into an observation group and a control group. In the observation group, 27 subjects received regular LQG-based breathing training for 12 weeks, while 20 subjects in the control group did not receive any intervention. The spine measuring instrument Spinal Mouse was adopted to detect the curvature and ROM of the thoracic and lumbar spines. Results: In the observation group, the changes in the curvatures of thoracic spine in the upright and forward-bending positions showed statistical significance after LQG exercise (P<0.05), while there was no notable difference in the control group. The comparison of ROM of the thoracic spine from the upright position to the forward-bending and load-bearing positions showed statistically significant difference in the observation group (P<0.05), while there was no significant difference in the control group. There was no significant difference in the lumbar curvature and ROM in the two groups after LQG exercise. Conclusion: LQG-based breathing training improves the curvature and ROM of the thoracic spine.

Objective:To explore the role of SUMOylaiton of peroxisome proliferator-activated receptor γ (PPARγ) in diabetes mellitus prompted inflammation and atherosclerosis in vascular and endothelial cells.Methods:From September 2014 to January 2017, 32 Sprague-Dawley rats in 14 weeks-old were divided into sham operated group, artery injured without diabetes group, artery injured with diabetes group and ubiquitin-conjugating enzyme 9 (UBC9) transfection group (Group D) by random digits table method with 8 rats each. Model of type 1 diabetes mellitus (T1DM) and rat carotid artery balloon injury was made in the assigned group. One rat was excluded because of model failure in each group. Systolic and diastolic common carotid artery diameter and intimal thickness of injured and healthy common carotid artery were evaluated by vascular ultrasound, and the standardized common carotid artery diastolic diameter (sCADD) was calculated. Histological tests and immunohistochemical staining were performed to evaluate intimal hyperplasia, and the ratio of intimal area to media area was calculated when the media area was equal. Human umbilical vein endothelial cells (HUVEC) were cultured 24 h in high glucose medium with different duration and concentration, and the expression levels of interleukin (IL)-8 and IL-1β mRNA were determined by real time reverse transcription polymerase chain reaction (RT-PCR), the expression level of UBC9 was determined by Western blot method, SUMOylation assay kit was used to evaluate SUMOylation of PPARγ. HUVEC was cultured in vitro and PPAR was stimulated by high glucose at different concentrations and different times PPARγ SUMOylation level. UBC9 was overexpressed by lentivirus in vivo and in vitro, and the PPARγ SUMOylation level was detected.Results:The intimal thickness, intimal area and ratio of intimal area to media area 8 weeks after carotid artery injuring in sham operated group, artery injured without diabetes group and artery injured with diabetes group were increased respectively: (0.026 ± 0.018), (0.084 ± 0.007) and (0.264 ± 0.022) mm; (0.18 ± 0.09) × 10 6, (0.32 ± 0.06) × 10 6 and (1.64 ± 0.22)×10 6 μm 2; 0.345 ± 0.073, 0.570 ± 0.080 and 2.710 ± 0.220, the sCADD was decreased respectively: 0.903 ± 0.084, 0.800 ± 0.071 and 0.330 ± 0.036, and there were statistical differences ( F = 10.40, 9.40, 8.20 and 8.60; P<0.05). After HUVEC was cultured in high glucose for 24 h, the IL-8 mRNA at sugar concentrations of 10, 20 and 40 mmol/L was 1.00 ± 0.11, 3.57 ± 0.22 and 4.07 ± 0.40, the IL-1β mRNA was 1.00 ± 0.07, 3.32 ± 0.29 and 5.13 ± 0.19, and there were statistical differences ( F = 73.05 and 205.80, P<0.05). The level of PPARγ SUMOylation and UBC9 in artery injured with diabetes group were significantly lower than those in artery injured without diabetes group (0.46 ± 0.25 vs. 1.00 ± 0.21 and 0.45 ± 0.02 vs. 1.00 ± 0.07), and there were statistical differences ( P<0.05); there was no statistical difference in PPARγ between 2 groups (0.94 ± 0.07 vs. 1.00 ± 0.04, P>0.05). The UBC9 and PPARγ SUMOylation at sugar concentrations of 0, 10, 20 and 40 mmol/L were decreased respectively (0.99 ± 0.05, 0.80 ± 0.06 and 0.62 ± 0.05; 1.00 ± 0.05, 0.57 ± 0.13 and 0.55 ± 0.08), and there were statistical differences ( F = 21.02 and 14.31, P<0.05); there was no statistical difference in PPARγ (1.00 ± 0.03, 0.90 ± 0.04 and 0.91 ± 0.05; F = 3.11, P>0.05). In HUVEC cultured in high glucose medium (20 mmol/L) for 6, 12, 24 and 48 h, the UBC9 and PPARγ SUMOylation were downregulated progressively (1.00 ± 0.09, 0.75 ± 0.05, 0.70 ± 0.08, 0.38 ± 0.04 and 0.35 ± 0.03; 1.00 ± 0.03, 0.86 ± 0.01, 0.59 ± 0.01, 0.51 ± 0.11 and 0.35 ± 0.08), and there were statistical differences ( F = 36.06 and 33.13, P<0.05); but there was no statistical difference in PPARγ (1.00 ± 0.03, 1.14 ± 0.02, 1.18 ± 0.17, 0.98 ± 0.01 and 1.04 ± 0.05; F = 1.90, P>0.05). After overexpression of UBC9 in rats with diabetes, histological analysis showed that UBC9 in artery injured without diabetes group, artery injured with diabetes group and UBC9 transfection group was 1.53 ± 0.18, 1.00 ± 0.22 and 3.62 ± 0.35, there was statistical difference ( F = 5.64, P<0.05). Ultrasonic test results show that in artery injured without diabetes group, artery injured with diabetes group and UBC9 transfection group intimal thickness was increased respectively: (0.077 ± 0.015), (0.216 ± 0.007) and (0.125 ± 0.014) mm, and there was statistical difference ( F = 27.18, P<0.05). Histological analysis showed that intimal area in artery injured without diabetes group, artery injured with diabetes group and UBC9 transfection group was (0.335 ± 0.066) ×10 6, (1.053 ± 0.103) ×10 6 and (0.544 ± 0.040) ×10 6 μm 2, the ratio of intimal area to media area was 0.63 ± 0.063, 2.03 ± 0.052 and 0.93 ± 0.100, there were statistical differences ( F = 13.58 and 53.96, P<0.05). Conclusions:Diabetes mellitus could inhibit the PPARγ SUMOylaiton and prompt inflammation and atherosclerosis in vascular and endothelial cells. Upregulation of PPARγ SUMOylaiton though UBC9 overexpressioncould play a protecting role in diabetes mellitus prompted atherosclerosis.

To achieve uniform soluble expression of multiple proteins in the same Escherichia coli strain, and simplify the process steps of antigen production in genetic engineering subunit multivalent vaccine, we co-expressed three avian virus proteins including the fowl adenovirus serotype 4 (FAdV-4) Fiber-2 protein, infectious bursal disease virus (IBDV) VP2 protein and egg-drop syndrome virus (EDSV) Fiber protein in E. coli BL21(DE3) cells after optimization of gene codon, promoter, and tandem expression order. The purified proteins were analyzed by Western blotting and agar gel precipitation (AGP). The content of the three proteins were well-proportioned after co-expression and the purity of the purified proteins were more than 80%. Western blotting analysis and AGP experiment results show that all the three co-expression proteins had immunoreactivity and antigenicity. It is the first time to achieve the three different avian virus antigens co-expression and co-purification, which simplified the process of antigen production and laid a foundation for the development of genetic engineering subunit multivalent vaccine.
Animals ; Antigens, Viral/genetics* ; Biological Assay ; Chickens/immunology* ; Escherichia coli/genetics* ; Infectious bursal disease virus/immunology* ; Poultry Diseases ; Vaccines, Synthetic/isolation & purification* ; Viral Structural Proteins/immunology* ; Viral Vaccines/immunology*

Objective To analyze the clinical characteristics and prognosis of patients with rectal gastrointestinal stromal tumor ( GIST) . Methods We collected the data of 31 rectal GIST patients definitely demonstrated by pathology and immunohistochemistry in the First Affiliated Hospital of Soochow University, the Second Affiliated Hospital of Soochow University, and Suzhou Wuzhong People′s Hospital from January 2008 to December 2016. The clinical characteristics, therapeutic modalities, and prognoses were retrospectively analyzed. Results Among the 31 rectal GIST patients, 16 underwent local resection and 15 underwent radical surgery. There was no significant difference on the three-year survival rate between the two groups [ 93. 8％ ( 15/16 ) VS 73. 3％ ( 11/15 ) , P=0. 135 ] . There was no significant difference on the survival rate between the oral imatinib patients and non-oral imatinib patients in the local resection group [ 75. 0％ ( 3/4) VS 100. 0％ ( 12/12) , P=0. 083] and the radical surgery group[ 77. 8％ ( 7/9) VS 66. 7％(4/6), P=0. 579]. The postoperative recurrence and metastasis rate of the two groups was no significantly different[31. 3％ (5/16) VS 53. 3％ (8/15), P=0. 213]. Conclusion The choice of surgical procedure has no significant effects on the survival rate and postoperative recurrence for rectal GIST patients, and whether oral imatinib or not has no significant effects on the survival of patients.

Objective To investigate the clinicopathological characteristics and the expression of related molecular markers of heterotopic pancreas for better understanding and avoiding overtreatment of this disease.Methods From 24th March 2009 to 10th November 2016,92 patients with heterotopic pancreas in upper digestive tract diagnosed after endoscopic submucosal dissection(ESD),were collected. Tissues were sectioned and pathologically classified by Heinrich classification.The expressions of seven different molecular markers including cytokeratin(CK)19,insulin,trypsin,Ki-67,p53,CD133 and CD56 were detected by immunohistochemistry staining. Clinical features, pathological features and immunohistochemical results were retrospectively analyzed.Analysis of variance and Kruskal-Wallis test were used.Results According to Heinrich classification,the percentages of type Ⅰ,Ⅱ,and Ⅲ of heterotopic pancreas were 27.2%(25/92),63.0%(58/92)and 9.8%(9/92),respectively.There was no type Ⅳ patients.The heterotopic pancreas mainly located in stomach with proportion being about 91.3%(84/92)of the total heterotopic pancreas.CK19(the marker of pancreas ducts),insulin(as marker of endocrine function)and trypsin(as the marker of exocrine function)were all expressed in heterotopic pancreas.The positive rate of CD56,a pancreatic neuroendocrine marker,was 66.3%(61/92).The umbilicus like depression was the typical endoscopic appearance of heterotopic pancreas,which only found in 29 patients(31.5%).The average rate of Ki-67,cell proliferation index,was(2.08 ± 1.41)%.The expression of mutant p53 was negative in all 92 cases of heterotopic pancreas.The average staining area of CD133,a marker of pancreatic cancer stem cell,was(2.53 ± 2.43)%.The average follow-up period of 92 patients was(43.6 ± 27.5)months.No relapse and malignant change were found and all patients survived after ESD.Conclusions Heterotopic pancreas has normal pancreatic construction and function.The cell proliferation index is low in heterotopic pancreas and no mutant p53 expression is found.The expression of CD133 is also low.Heterotopic pancreas is a congenital benign disease which requires a long-term follow-up rather than overtreatment.

Coronary atherosclerosis (CAS) has been recognized as a chronic inflammatory disease in recent years.As an important component of innate immunity,complement system plays an important role in the pathogenesis of CAS.Multiple components of plaque can activate complement proteins,and the activation of complement promotes the progression of plaque lesions in turn.However,recent studies have also found that some proteins of the complement system may also paly an atheroprotective role in the early stage of atherosclerosis.In addition to the direct effects,complement system is also involved in CAS-related pathological changes.Complement proteins may be promising biomarkers,as confirmed by clinical studies which suggest that circulation levels of some complement proteins can predict the risk and prognosis of CAS.This review mainly introduced the origin and activation of complement molecules inside the plaques,their important roles in the pathogenesis of CAS,and the value of complement molecules as potential predictors in clinical.

OBJECTIVETo explore the related factors of optical coherence tomography (OCT) detected in-stent heterogeneous neointimal in coronary stents.

METHODSA total of 143 cases of coronary heart disease patients with OCT detected in-stent neointimal hyperplasia in Fuwai hospital from September 2009 to April 2012 were included in this study and patients data were retrospectively analyzed. Patients were divided into heterogeneous intima group(26 cases) and homogeneous intima group(117 cases)according to neointimal characteristics. Clinical features and OCT characteristics of the 2 groups were compared and binary logistic regression analysis was performed to analyze the risk factors of in-stent heterogonous neointimal hyperplasia.

RESULTSCompared to homogeneous intima group, patients in heterogeneous intima group had significantly higher cholesterol level ((5.31±1.11)mmol/L vs.(4.70±0.94)mmol/L, P=0.005), low-density lipoprotein cholesterol level ((2.57±0.87)mmol/L vs.(2.29±0.46)mmol/L, P=0.021) and triglyceride level (2.12(1.82-2.87)mmol/L vs. 1.90(1.73-2.11)mmol/L, P=0.015). Moreover, the percent of percutaneous coronary intervention (PCI) because of acute coronary syndrome (23.1%(6/26) vs. 6.8%(8/117), P=0.022) and the thin cap neoatheroma (5.8%(28/481)vs. 3.9%(89/2 276), P=0.043) were also significantly higher in heterogeneous intima group than in homogeneous intima group. Binary logistic regression analysis showed that low-density lipoprotein cholesterol (OR=2.74, 95%CI 1.04-7.24, P=0.042), triglyceride (OR=2.88, 95%CI 1.05-7.89, P=0.040), PCI for acute coronary syndrome (OR=12.74, 95%CI 2.69-60.49, P=0.001), and cerebrovascular disease (OR=13.09, 95%CI 2.16-79.53, P=0.005) were risk factors for in-stent heterogenous intima. Time post stent implantation was protective factor for in-stent heterogenous intima (OR=0.63, 95%CI 0.42-0.96, P=0.033).

CONCLUSIONOCT detected heterogeneous intima is correlated with level of blood lipid, PCI for acute coronary syndrome and history of cerebrovascular disease, and it may lead to unstable intima.

Acute Coronary Syndrome ; Coronary Artery Disease ; Humans ; Hyperplasia ; Neointima ; Percutaneous Coronary Intervention ; Retrospective Studies ; Risk Factors ; Stents ; Tomography, Optical Coherence ; Tunica Intima