Objective To investigate the effect of microRNA-146a-5p(miR-146a-5p)on high glucose-induced injury of renal tubular epithelial cells by regulating high mobility group A2(HMGA2).Methods Human renal tubular epithelial cells(HK-2)were cultured in vitro and di-vided into different groups according to various treatment methods:normal control group(Con group),high glucose group(HG group),HG+miR-146a-5p group,HG+si-HMGA2 group,HG+miR-146a-5p+pcDNA-HMGA2 group,miR-NC group,miR-146a-5p group,anti-miR-NC group,and anti-miR-146a-5p group.Real-time quantitative polymerase chain reaction(qRT-PCR)was used to detect the expression levels of miR-146a-5p and HMGA2 mRNA in each group.A dual-luciferase reporter assay was employed to verify the targeting relationship between miR-146a-5p and HMGA2.Western blot was utilized to detect the expression levels of cleaved-caspase3,cleaved-caspase9,and HMGA2 protein in each group.Flow cytometry was applied to determine the apoptosis rate of cells in each group.Enzyme-linked immunosorbent assay(ELISA)was used to detect the levels of inflam-matory factors[interleukin-6(IL-6),tumor necrosis factor-α(TNF-α),interleukin-8(IL-8)]in each group.The levels of oxidative stress indicators[catalase(CAT),superoxide dismutase(SOD),malondialdehyde(MDA)]in each group were also measured.Results A nucleotide se-quence complementary to miR-146a-5p was found in the 3'-untranslated region(3'-UTR)of HMGA2.The expression level of HMGA2 protein in the miR-146a-5p group was lower than that in the miR-NC group,while was higher in the anti-miR-146a-5p group than that in the anti-miR-NC group(P＜0.05).The total apoptosis rate,early apoptosis rate,late apoptosis rate,and the levels of cleaved-caspase3 protein,cleaved-caspase9 protein,HMGA2 mRNA,HMGA2 protein,TNF-α,IL-6,IL-8,and MDA in the HG group were higher than those in the Con group,whereas the levels of miR-146a-5p,CAT,and SOD were lower(P＜0.05).In the HG+miR-146a-5p group,these indicators showed opposite trends compared to the HG group(P＜0.05).Similarly,in the HG+si-HMGA2 group,the total apoptosis rate,early apoptosis rate,late apoptosis rate,and the levels of cleaved-caspase3 protein,cleaved-caspase9 protein,HMGA2 mRNA,HMGA2 protein,TNF-α,IL-6,IL-8,and MDA were lower than those in the HG group,while the levels of CAT and SOD were higher(P＜0.05).In the HG+miR-146a-5p+pcDNA-HMGA2 group,the trends of these indicators were consistent with those between the HG group and the HG+miR-146a-5p group,showing statistically significant differences when compared to the HG+miR-146a-5p group(P＜0.05).Conclusion MiR-146a-5p may play a regulatory role in high glucose-induced injury of re-nal tubular epithelial cells by targeting HMGA2.

Objective To study the effect and molecular mechanism of vitamin D(VitD)on Tfh cells of MRL/lpr lupus mice.Methods C57/B6 mice and MRL/lpr lupus mice were transfected with siRNA to construct VDR knockout mouse models.Splenic Tfh cells of C57/B6 mice and MRL/lpr lupus mice were divided into control group,lupus group and VDRsiRNA lupus group(treated with vitamin D 0,1 and 10 nmol·L-1,respectively)by siRNA transfection.The percentage of Tfh cells was detected by flow cytometry.MRL/lpr lupus mice Peyer node Tfh cells were randomly divided into 7 groups,blank control group,vitamin D dose groups of 1 and 10 nmol·L-1,paricalcitol group(VitD 10 nmol·L-1+PA),VDRsiRNA control group,VDRsiRNA group(VitD 10 nmol·L-1),CaN inhibitor group(VitD 10 nmol·L-1+CsA),and incubated for 72 h.The concentration of calcium ions in Tfh of each group was detected.The expressions of AT1R,NFAT,CaN and P-CaN in Tfh cells were determined by Western blotting.Results The percentage of Tfh cells decreased significantly with the increase of vitamin D dose.Vitamin D CaN reduce the intracellular calcium concentration of Tfh,up-regulate the expression of AT1 protein in Tfh cells,and down-regulate the expression of CAN,P-CaN and NFAT protein in a dose-dependent manner,and the effect is more obvious when combined with PA.Conclusion Vitamin D may regulate the activation of follicular T helper cells in MRL/lpr mice via the Ca-CaN-NFAT pathway.

Objective To study the effect and molecular mechanism of vitamin D(VitD)on Tfh cells of MRL/lpr lupus mice.Methods C57/B6 mice and MRL/lpr lupus mice were transfected with siRNA to construct VDR knockout mouse models.Splenic Tfh cells of C57/B6 mice and MRL/lpr lupus mice were divided into control group,lupus group and VDRsiRNA lupus group(treated with vitamin D 0,1 and 10 nmol·L-1,respectively)by siRNA transfection.The percentage of Tfh cells was detected by flow cytometry.MRL/lpr lupus mice Peyer node Tfh cells were randomly divided into 7 groups,blank control group,vitamin D dose groups of 1 and 10 nmol·L-1,paricalcitol group(VitD 10 nmol·L-1+PA),VDRsiRNA control group,VDRsiRNA group(VitD 10 nmol·L-1),CaN inhibitor group(VitD 10 nmol·L-1+CsA),and incubated for 72 h.The concentration of calcium ions in Tfh of each group was detected.The expressions of AT1R,NFAT,CaN and P-CaN in Tfh cells were determined by Western blotting.Results The percentage of Tfh cells decreased significantly with the increase of vitamin D dose.Vitamin D CaN reduce the intracellular calcium concentration of Tfh,up-regulate the expression of AT1 protein in Tfh cells,and down-regulate the expression of CAN,P-CaN and NFAT protein in a dose-dependent manner,and the effect is more obvious when combined with PA.Conclusion Vitamin D may regulate the activation of follicular T helper cells in MRL/lpr mice via the Ca-CaN-NFAT pathway.

Objective:To design a lung nodule detection algorithm based on the improved YOLOv7 model.Methods:Firstly, in the PAFPN structure, a lightweight upsampling operator CARAFE is introduced to improve the lung nodule detection accuracy. Then an enhanced small-scale detection layer is added to enhance the detection performance for small-target lung nodules, while the number of trained parameters can be reduced and the model complexity can be lowered. An enhanced small-scale detection layer is added to the YOLOv5 model algorithm while comparing it with the original YOLOv5 model algorithm, the original YOLOv7 model algorithm, and the improved YOLOv7 model algorithm in terms of the total loss of the training set of the improvement points, while ensuring that the parameter indexes remain unchanged. The original YOLOv7 model algorithm and the improved YOLOv7 model algorithm are used to process the 2 test set images and compare them with other classical lung nodule detection algorithms Mask R-CNN, YOLOv3, YOLOv5s and YOLOv7.Results:Compared with the original YOLOv5 model algorithm, the improved YOLOv5 model algorithm with the addition of an enhanced small-scale detection layer has a 1.3% increase in precision, a 3.5% increase in recall, a 3.1% increase in mean average precision (mAP), a 25.3% decrease in parameters amount, and a decrease in the complexity of the network; whereas the improved YOLOv7 model algorithm has a 1.8% increase in mAP, a 28.3% decrease in parameters amount, and the model complexity decreased by 5 G. Adding the enhanced small-scale detection layer with replacement of the special diagnostic fusion network as a lightweight up-sampling operator CARAFE algorithm can minimize the total loss of the model during the training process. The original YOLOv7 model algorithm is more accurate but still has missed detections and false positives. When reasoning about image 1, the original YOLOv7 model has a missed detection. And when reasoning about image 2, the original YOLOv7 model has a false positive. The improved YOLOv7 model is well improved in both missed detection and false positives. Compared with the classical model algorithm, the precision, recall, and mAP of the improved YOLOv7 model algorithm were 91.7%, 89.1%, and 93.5%, respectively.Conclusions:The improved YOLOv7 model has stronger feature expression ability and uses fewer parameters, which can effectively improve the detection precision of lung nodules.

Objective : To explore the expression of chemokine CXCL1 in proliferative infantile hemangioma (IH) , and to study the effect of exogenous CXCL1 on hemangioma stem cells (HemSCs) . Methods : Immunohistochemistry was used to explore the expression of CXCL1 in proliferative IH specimens.Primary HemSCs were isolated from IH tissues by CD133 magnetic beads.5 groups of CXCL1 with different concentrations(0,10,20,50 and 100 ng/ml) were co-cultured with HemSCs, and the effects of exogenous CXCL1 on HemSCs were studied by cell viability and migration experiments. Results : CXCL1 was expressed in the interstitial tissues of proliferative IH.The overall expression of CXCL1 in proliferative IH was low, but the expression of CXCL1 in the proliferative IH interstitial tissues was higher than that of the adjacent interstitial tissues.The CXCL1 positive area rate was(0.773±0.101)% in the tumor compared with(0.268±0.081)% in the adjacent tumor, and the difference was statistically significant(t=7.843,P<0.001).Exogenous CXCL1 promoted the proliferation of HemSCs, and there were statistical differences after adding different concentrations of CXCL1 to HemSCs for 24,48,and 72 h(F=14.610,P<0.001;F=14.430,P<0.001;F=5.388,P<0.01).But the exogenous CXCL1 did not affect the migration ability of HemSCs. Conclusion The expression of CXCL1 in proliferative IH interstitial tissues is higher than that in adjacent interstitial tissues, and exogenous CXCL1 promotes the proliferation of HemSCs.

Klebsiella pneumoniae(K.pneumoniae)is an important pathogen that can cause severe hospital-and community-acquired infections.To systematically investigate its methylation features,we determined the whole-genome sequences of 14 K.pneumoniae strains covering varying serotypes,multilocus sequence types,clonal groups,viscosity/virulence,and drug resistance.Their methy-lomes were further characterized using Pacific Biosciences single-molecule real-time and bisulfite technologies.We identified 15 methylation motifs[13 N6-methyladenine(6mA)and two 5-methylcytosine(5mC)motifs],among which eight were novel.Their corresponding DNA methyl-transferases were also validated.Additionally,we analyzed the genomic distribution of GATC and CCWGG methylation motifs shared by all strains,and identified differential distribution pat-terns of some hemi-/un-methylated GATC motifs,which tend to be located within intergenic regions(IGRs).Specifically,we characterized the in vivo methylation kinetics at single-base resolu-tion on a genome-wide scale by simulating the dynamic processes of replication-mediated passive demethylation and MTase-catalyzed re-methylation.The slow methylation of the GATC motifs in the replication origin(oriC)regions and IGRs implicates the epigenetic regulation of replication initiation and transcription.Our findings illustrate the first comprehensive dynamic methylome map of K.pneumoniae at single-base resolution,and provide a useful reference to better understand epigenetic regulation in this and other bacterial species.

The rapid spread of carbapenemase-producing Klebsiella pneumoniae(cpKP)poses seri-ous threats to public health;however,the underlying genetic basis for its dissemination is still unknown.We conducted a comprehensive genomic epidemiology analysis on 420 cpKP isolates col-lected from 70 hospitals in 24 provinces/autonomous regions/municipalities of China during 2009-2017 by short-/long-read sequencing.The results showed that most cpKP isolates were categorized into clonal group 258(CG258),in which ST11 was the dominant clone.Phylogenetic analysis revealed three major clades including the top one of Clade 3 for CG258 cpKP isolates.Additionally,carbapenemase gene analysis indicated that blaKPC was dominant in the cpKP isolates,and most blaKPC genes were located in five major incompatibility(Inc)groups of blaKPC-harboring plasmids.Importantly,three advantageous combinations of host-blaKPC-carrying plasmid(Clade 3.1+3.2-IncFⅡpHN7A8,Clade 3.1+3.2-IncFⅡpHN7A8:IncR,and Clade 3.3-IncFⅡpHN7A8:InCpA1763-KPC)were identified to confer cpKP isolates the advantages in both genotypes(strong correlation/coevolution)and phenotypes(resistance/growth/competition)to facilitate the nationwide spread of ST11/CG258 cpKP.Intriguingly,Bayesian skyline analysis illustrated that the three advanta-geous combinations might be directly associated with the strong population expansion during 2007-2008 and subsequent maintenance of the population of ST11/CG258 cpKP after 2008.We then examined drug resistance profiles of these cpKP isolates and proposed combination treatment regimens for CG258/non-CG258 cpKP infections.Thus,the findings of our systematical analysis shed light on the molecular epidemiology and genetic basis for the dissemination of ST11/CG258 cpKP in China,and much emphasis should be given to the close monitoring of advantageous cpKP-plasmid combinations.

Objective: To report the effects of operative treatment of Sneppen Ⅴ talus fracture through the approach for malleolus medialis Ⅴ osteotomy plus hollow compression screw fixation. Methods: From January 2015 to January 2019, 16 patients with Sneppen Ⅴ talus fracture were treated at Department Ⅱ of Hand & Foot Microsurgery, The Second Affiliated Hospital to Inner Mongolia Medical University. They were 14 men and 2 women with a mean age of 38.4 years (range, from 20 to 55 years). All fractures were fixed with hollow compression screws through the approach for malleolus medialis Ⅴ osteotomy. The ankle and hindfoot functional scoring system developed by American Orthopaedic Foot and Ankle Society (AOFAS) was used to evaluate the clinical outcomes. Results: All patients were followed up for a mean time of 12.6 months (range, from 6 to 30 months). The mean operation time was 68.4 minutes (range, from 52 to 96 minutes); the mean amount of hemorrhage during operation was 96.8 mL (range, from 48 to 122 mL); the mean period of bone union was 4.8 months (range, from 3 to 8 months). The postoperative mean AOFAS score was 75.3 points (range, from 43 to 91 points). Complications occurred in 4 cases, including one case of talus ischemic necrosis, one case of partial talus ischemic necrosis accompanied by tibial arthritis, one case of subtalar arthritis, and one case of combined tibial, talar and subtalar arthritis. All incisions obtained primary healing, with no complications like infection, screw breakage, delayed union or nonunion. Conclusion Operative treatment of Sneppen Ⅴ talus fracture through the approach for malleolus medialis Ⅴ osteotomy plus hollow compression screw fixation can provide sufficient operative exposure to facilitate reduction and fixation of the talus fracture so that the ischemic necrosis of the talus and traumatic arthritis can be effectively reduced.

Objective To study the transcriptional regulation of vp1667 by H-NS in Vibrio parahaemolyticus.Methods Total RNAs were extracted from Δhns and WT strains.Quantitative RT-PCR was carried out to calculate the transcriptional variation of vp1667 between Δhns and WT.Primer extension assay was also employed to detect the transcription start site and the promoter activity (i.e.the amount of primer extension products) of vp1667 in Δhns and that in WT.The promoter DNA region of vp1667 was amplified, purified, and cloned into the corresponding restriction endonuclease sites of pHRP309 that harbors a gentamicin resistance gene and a promoterless lacZ reporter gene.The recombinant pHRP309 plasmid was transformed into Δhns and WT, respectively, while β-galactosidase activity in cellular extracts was measured using a β-galactosidase enzyme assay system.The over-expressed His-H-NS was purified under native conditions with nickel loaded HiTrap Chelating Sepharose columns.The electrophoretic mobility shift assay (EMSA) and DNaseⅠ footprinting were then applied to analyze the DNA-binding activity of His-H-NS to vp1667 promoter region in vitro.Results and Conclusion The primer extension assay detected one transcription start site for vp1667, which was located at 28 bp upstream of vp1667, and its transcribed activity was under the negative control of the H-NS.The EMSA and DNaseⅠ footprinting assay results showed that His-H-NS was unable to bind to the promoter-proximal DNA region of vp1667, suggesting that H-NS indirectly inhibits the transcription of vp1667.

Objective To characterize the whole-sequence of plasmid pB557-NDM isolate from Enterbacter cloacae and elaborate its antibiotic-resistant mechanisms .Methods Antibiotic resistance genes were determined by PCR , followed by amplicon sequencing .The activity of class A/B/D carbapenemases was determined by modified Carba NP test .Conjugation experiments were performed to verify the transferability of plasmid pB 557-NDM.The minimum inhibitory concentration (MIC) values of bacterial strains were tested using VITEK 2.The genetic structure, mobile elements and antibiotic-resistant mechanisms of transferable plasmid pB 557-NDM were determined by a whole genome sequencing method .Results The modified CarbaNP test showed that B557 and B557-EC600 had class B carbapenemase activity , and that the blaNDM was carried by plasmid pB557-NDM.This plasmid could be transferred into E.coli through conjugation experiments and therefore could confer corresponding antibiotic resistances to the transconjugant B 557-EC600.Plasmid pB557-NDM was an IncA/C2 plasmid, whose total length was 141.65 kb, and the GenBank accession number was KX786648.It had two inserted regions.One was the blaCMY-6 region where the blaCMY gene was carried by a transposition unit IS Ecp1-blaCMY , the other was the blaNDM-1 region which consisted of a ΔTn1696-In46-rmtC-ISKpn14-ΔTn125 complex structure.Conclusion The production of plasmid pB557-NDM in strain B557 contributes most to its high resistance to many antibiotics .The blaNDM-1 gene is carried in a trancated transposition ΔTn125.