Objective:To evaluate the role of the sodium leak channel (NALCN) in the hippocampal dentate gyrus (DG) in the social behavior of mice.Methods:Thirty-nine male wild-type C57BL/6 mice, aged 6-8 weeks, weighing 18-22 g, were used in this study. Three mice were sacrificed to verify the expression and co-expression of NALCN with neuronal nuclear antigen (NeuN) in the hippocampal DG using the immunofluorescent staining. The remaining 36 mice were divided into 2 groups ( n=18 each) by the random number table method: control group (group C) and NALCN gene knockdown group (group KO). NALCN-shRNA virus was injected in group KO, and scrambled-shRNA virus was injected in group C. The three box social test and open field test were performed at 3 weeks after the virus injection. Mice were sacrificed under anesthesia after the behavioral test, hippocampal tissues were collected, and the injection location of the virus was verified with a fluorescence microscope, and the NALCN protein and mRNA expression in the hippocampal DG was detected by Western blot and real-time polymerase chain reaction, respectively. Results:NALCN and NeuN co-expressed a lot on the same neuron in the hippocampal DG of mice, indicating that NALCN was widely expressed on the neurons in the hippocampal DG. Compared with group C, the expression of NALCN and mRNA in the hippocampal DG was significantly down-regulated, and the social novelty preference disappeared ( P<0.05), and no significant change was found in the social ability and each parameter in the open field test in group KO ( P>0.05). Conclusions:NALCN in the hippocampal DG is involved in the regulation of social memory in mice, and the down-regulated expression of NALCN can lead to the loss of social novelty preference in mice.

Objective:To explore the effects of mechanical factors on the morphology of the growth plate and proximal femur development, in order to provide a mechanical basis for the mechanism of Cam lesions in femoral acetabular impingement (FAI).Methods:Using CT scan data of hip joint from adolescents, we constructed three-dimensional finite element models of normal and extended proximal femoral epiphyseal growth plates. Dynamic mechanics and position data from daily activities (walk, stand on one and two feet, go upstairs and downstairs, sit and stand up, squat and stand up) and basketball layup drills were collected from five healthy volunteers using the Vicon system. The mean values of femoral head mechanical loads and positions at peak values, movement onset, and termination were selected for finite element analysis to observe the distribution of equivalent stress, normal stress, and shear stress on the two growth plate models under different activity modes.Results:Successful construction of three-dimensional finite element models for normal and extended proximal femoral epiphyseal growth plates was achieved. Mechanical load values and spatial positions of the femoral head during daily activities and layup movements were obtained. In daily activities, peak equivalent stress values for normal and extended growth plates ranged from 1.6 to 11.0 MPa; compressive stress ranged from 1.7 to 12.0 MPa; tensile stress ranged from 0.5 to 10.0 MPa; and shear force ranged from 0.4 to 7.1 MPa. The compressive stress during walking, standing on one or both feet, going up and down stairs, sitting and standing, and squatting was concentrated in the central and lateral areas; tensile stress was concentrated in the central and medial areas; shear force was concentrated in the medial area and the anterior and posterior edges of the growth plate. During a layup, the compressive stress in the center, outer upper side, and medial edge of the growth plate of the normal model was notably concentrated, with peaks fluctuating between 5.5-19.0 MPa, 5.7-11.0 MPa, and 5.4-7.3 MPa respectively; tensile stress and shear force were concentrated at the inner and outer edges as well as in the central area, with peaks fluctuating between 3.0-24.0 MPa and 3.0-26.0 MPa respectively, these values were significantly different from those observed during daily activities. For the elongated growth plate, compressive stress was concentrated at the central and lateral edges with peaks fluctuating between 17.0-41.0 MPa and 17.0-38.0 MPa respectively; simulated shear stress and tensile stress showed significant concentration at peaks fluctuating between 4.9-34.0 MPa, also significantly different from those observed during daily activities.Conclusion:The difference in mechanical distribution between daily activities, basketball layup training, as well as between normal and extended growth plates may be the mechanical initiating factor in the development of extended growth plates and the formation of Cam lesions, it may be the mechanical initiating factor for the development of extended growth plates and the collision of Cam lesions in FAI.

Objective:To evaluate the relationship between Na + leak current channel (NALCN) in hippocampal dentate gyrus (DG) region and cognitive function in mice. Methods:Thirty-nine male wild type C57BL/6 mice, aged 6-8 weeks, weighing 18-22 g, were studied.Three mice were sacrificed to verify the expression of NALCN co-localized with neuronal nuclear antigen (NeuN) in the hippocampal DG region through immunofluorescence technique.The remaining 36 mice were divided into 2 groups ( n=18 each) by the random number table method: control group (group C) and NALCN gene knockout group (group KO). NALCN-shRNA virus was injected in hippocampal DG region in group KO, and scrambled-shRNA virus was injected in group C. Three weeks after virus injection, behavioral tests (Y maze test and open field test) were performed, then the animals were sacrificed, and the hippocampal tissues were removed for determination of the expression of NALCN protein and mRNA using Western blot and real-time polymerase chain reaction. Results:NALCN and NeuN colocalized a lot on the same neuron in the hippocampal DG region of mice, and NALCN was widely expressed in the hippocampal DG region.Compared with group C, the expression of NALCN protein and mRNA was significantly down-regulated, the times of entering the new arm were reduced, the duration of staying at the new arm was shortened ( P<0.05), and no significant change was found in the parameters mentioned above in the open field test in group KO ( P>0.05). Conclusions:NALCN in the hippocampal DG region is involved in the regulation of cognitive function in mice, and the down-regulation of NALCN may lead to cognitive decline.

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 observe the effects of veno-venous extracorporeal membrane oxygenation (VV-ECMO)combined with prone position ventilation (PPV) on oxygenation index (PaO 2/FiO 2), respiratory compliance (Crs) and vasoactive inotropic score (VIS) in severe acute respiratory distress syndrome (ARDS) patients. Methods:Eighteen patients with severe ARDS requiring VV-ECMO support in Liuzhou People's Hospital from June 2018 to April 2020 were selected for retrospective analysis, and 8 patients among of these cases received PPV after VV-ECMO. The differences in PaO 2/FiO 2, VIS and Crs before and 1, 2 or 3 days after treatment were compared between VV-ECMO group and VV-ECMO combined with PPV group, as well as the differences in these indices before PPV and 2 hours after PPV daily in VV-ECMO combined with PPV group. The incidence of adverse events in two groups were also observed. Results:Before treatment, there was no significant difference in PaO 2/FiO 2, Crs between two groups. Over time, PaO 2/FiO 2 and Crs increased and VIS decreased in both groups. Compared with before treatment, there were statistically significant differences in PaO 2/FiO 2 and VIS from 1 day after treatment [PaO 2/FiO 2 (mmHg, 1 mmHg = 0.133 kPa): VV-ECMO group was 197.75±39.80 vs. 75.57±7.44, VV-ECMO combined with PPV group was 255.20±31.92 vs. 68.24±11.64; VIS: VV-ECMO group was 5.51±3.72 vs. 10.20±7.10, VV-ECMO combined with PPV group was 6.73±3.32 vs. 14.50±2.48, all P < 0.05], up to 3 days after treatment [PaO 2/FiO 2 (mmHg): VV-ECMO group was 231.96±32.76 vs. 75.57±7.44, VV-ECMO combined with PPV group was 285.61±19.40 vs. 68.24±11.64; VIS: VV-ECMO group was 2.26±1.90 vs. 10.20±7.10, VV-ECMO combined with PPV group was 2.13±1.55 vs. 14.50±2.48, all P < 0.05], and the PaO 2/FiO 2 1 day and 3 days after treatment in VV-ECMO combined with PPV group were significantly higher than those in VV-ECMO group (mmHg: after 1 day of treatment was 255.20±31.92 vs. 197.75±39.80, after 3 days of treatment was 285.61±19.40 vs. 231.96±32.76, both P < 0.05). Before treatment, Crs of VV-ECMO combined with PPV group was significantly lower than that of VV-ECMO group (mL/cmH 2O: 17.91±0.82 vs. 20.54±1.26, P < 0.05). From 1 day after treatment, the Crs in VV-ECMO combined with PPV group was significantly higher than that before treatment (mL/cmH 2O: 21.20±1.50 vs. 17.91±0.82), the peak value was (24.93±2.18) mL/cmH 2O on 3 days after treatment, however, there was no significant difference between the two groups (all P > 0.05). In VV-ECMO combined with PPV group, compared with before PPV treatment, the PaO 2/FiO 2 and Crs of 2 hours after PPV treatment in 1, 2 and 3 days were significantly rose, and it reached the highest level in 3 days [PaO 2/FiO 2(mmHg): 285.61±19.40 vs. 189.91±28.34, Crs (mL/cmH 2O): 24.93±2.18 vs. 23.35±1.45, both P < 0.05]; the VIS was only increased in 2 hours after PPV treatment on the first day than before (6.73±3.32 vs. 6.38±3.22, P < 0.05). There were no related serious adverse events happened after PPV treatment. Conclusions:The combination of PPV during VV-ECMO could further increase PaO 2/FiO 2, improve hypoxemia and implement further protective lung ventilation to reduce the potential hazards during mechanical ventilation. In addition, no serious adverse events were observed in this study, suggesting PPV is safe during VV-ECMO.

Objective:To systematically compare the efficacy of different drugs in alleviating remifentanil-induced hyperalgesia.Methods:Databases such as PubMed, Cochrane Library, EMBASE, Web of Science, CNKI, Wanfang Data and CBM were searched using computers from inception to May 2020.The randomized controlled trials comparing the efficacy of different intervention measures for alleviating remifentanil-induced hyperalgesia were searched.After independently identifying the literature, the two reviewers conducted data extraction and evaluated the bias of the included studies, and Stata 14.0, ADDIS 1.16.5 and R4.0.2 softwares were used to analyze the data.Results:Thirty randomized controlled trials were included in our study.Compared with placebo, 3 out of 6 drugs could alleviate remifentanil-induced hyperalgesia, and the probability order for the effect was as follows: butorphanol with MD value (95% CI)-1.50 (-2.80, -0.24), dexmedetomidine with MD value (95% CI)-1.20 (-2.40, -0.09) and ketamine with MD value (95% CI) -0.88 (-1.60, -0.16). After sensitivity analysis, the efficacy of butorphanol remained to be verified.Two drugs could decrease the dosage of opioids within 24 h after operation, and the probability order for the effect was as follows: dexmedetomidine with MD value (95% CI) -14.00 (-28.00, -0.19) and ketamine with MD value (95% CI) -9.20 (-18.00, -0.08). One drug could decrease the incidence of postoperative nausea and vomiting within 24 h after operation: dexmedetomidine with RR value (95%CI) 0.28 (0.16, 0.22). Conclusion:The results of network meta-analyses show that dexmedetomidine has the best efficacy in alleviating remifentanil-induced hyperalgesia.

Objective:To analyze the efficacy of dienogest combined with traditional Chinese medicine(TCM) in the treatment of endometriosis (EMs) and its influence on pregnancy and recurrence.Methods:A total of177 patients with EMs admitted to Affiliated Hospital of Shandong University of Chinese Medicine and Linyi Hospital of Traditional Chinese Medicine from December 2018 to June 2019 were divided into the observation group (88 cases) and the control group (89 cases) according to the random number table method. The control group was treated with dienogest 25 mg once a day, and the observation group was treated with dienogest combined with TCM decoction. They all were treated for 6 months. The clinical efficacy, pregnancy and recurrence of the two groups were compared, and EMs health profile-5 (EHP- 5) of the Chinese version was used to evaluate patients′ quality of life of the two groups.Results:The total effective rate in the observation group was higher than that in the control group: 92.04%(81/88) vs. 79.78%(71/89), and the difference was statistically significant ( χ2 = 5.492, P<0.05). The time of ovulation recovery and menstrual recovery in the observation group were shorter than those in the control group: (13.38 ± 3.60) d vs. (14.69 ± 3.15) d, (26.48 ± 4.23) d vs. (28.72 ± 3.40) d, and the differences were statistically significant ( P<0.05). There was no statistically significant difference in the 1-year pregnancy rate between the two groups ( P>0.05). The 1-year recurrence rate in the observation group was lower than in the control group: 1.14%(1/88) vs. 10.11%(9/89), and the difference was statistically significant ( P<0.05). The scores of pain, emotional state, sense of control, sexual life, doctor-patient relationship , infertility, and treatment effect in EHP-5 in the observation group after the treatment were lower than those in the control group, and the differences were statistically significant ( P<0.05). Conclusions:Compared with dienogest treatment alone, the combination of TCM treatment can further improve the efficacy. It is beneficial to pregnancy. It can reduce the recurrence rate, improve the quality of life more significantly, and has great safety.

Objective:To evaluate the therapeutic effects and prognostic factors of neoadjuvant chemoradiotherapy for locally advanced esophageal squamous cell carcinoma.Methods:The clinical data of a total of 148 patients with locally advanced esophageal squamous cell carcinoma enrolled in the Affiliated Cancer Hospital of Zhengzhou University from 2007 to 2017 were retrospectively analyzed. The patients received 5-Fu/Cisplatin or Paclitaxel/Cisplatin for chemotherapies. The total treatment dose for the radiotherapy was delivered at 36-40Gy under conventional fractionation. Kaplan-Meier method was used to calculate survival rates, and Log-rank test and Cox model were performed for univariate analysis and multivariate analysis, respectively.Results:The overall survival (OS) rates of 1-, 3-and 5-year were 74%, 51% and 51%, respectively, with a median survival time (MST) of 72.4 months. The carcinoma/disease-free survival (DFS) rates for 1, 3, 5 years were 60%, 51%, 45%, respectively, with a median time of 60.1 months. The 1-, 3-and 5-year OS rates of the pCR group were 86%, 70%, 70%, the ones of which in the non-pCR group were 70%, 44%, 43%, respectively ( P=0.002). The 1-, 3-and 5-year DFS rates were 76%, 71%, 68% for the pCR group, and 53%, 43%, 37% for the non-pCR group, respectively ( P=0.002). In pN(-) group and pN(+ ) group, the 1-, 3-and 5-year OS rates were 83%, 56%, 55% and 50%, 38%, 38%( P=0.004), respectively. Further, the 1-, 3-and 5-year DFS rates were 66%, 56%, 51% for the pN(-) group, and 43%, 38%, 31% for the pN(+ ) group ( P=0.006), respectively. Multivariate analysis revealed that pCR and pN status were independent prognostic factors for OS and DFS ( P=0.012, 0.011 and P=0.025, 0.033). Conclusion:Neoadjuvant chemoradiotherapy demonstrated significant therapeutic effects in the treatment of locally advanced esophageal squamous cell carcinoma, while pCR and pN status served as independent prognostic factors.

Objective:To investigate the correlation between white matter hypertensities (WMHs) and the overall burden of cerebral small vessel disease (CSVD) and clinical outcome of patients with acute ischemic stroke.Methods:From November 2018 to June 2019, patients with acute ischemic stroke hospitalized in the Department of Neurology, the Affiliated Jiangning Hospital of Nanjing Medical University were enrolled prospectively. Their demographic and clinical data were collected. The National Institutes of Health Stroke Scale (NIHSS) was used to evaluate the baseline severity of stroke. The total burden of CSVD was evaluated according to the head MRI findings. The severity of WMHs was assessed based on Fazekas scale. The modified Rankin Scale (mRS) was used to evaluate the outcomes at 90 d after onset. The mRS score 0-1 was defined as good outcome, and ≥2 was defined as poor outcome. Multivariate logistic regression analysis was used to determine independent risk factors for poor outcomes. Results:A total of 153 consecutive patients with acute ischemic stroke were enrolled, of which 126 (82.35%) had a good outcome and 27 (17.65%) had a poor outcome. There were significant differences in age, baseline NIHSS score, lipoprotein-associated phospholipase A 2, total Fazekas score, periventricular WMHs score, and deep WMHs score between the two groups, while there was no significant difference in the total burden of CSVD. Multivariate logistic regression analysis revealed that the baseline NIHSS score (odds ratio 1.245, 95% confidence interval 1.023-1.515; P=0.028) and the total Fazekas score (odds ratio 1.635, 95% confidence interval 1.049-2.549; P=0.030) were the independent risk factors for poor outcomes at 90 d after the onset in patients acute ischemic stroke. Conclusions:The overall burden of CSVD is not associated with the short-term outcomes in patients with acute ischemic stroke. WMHs and baseline NIHSS score are the independent risk factors for poor short-term outcomes in patients with acute ischemic stroke.