Induce male sterility by CRISPR/Cas9-mediated mitochondrial genome editing in tobacco.

Genome editing has become more and more popular in animal and plant systems following the emergence of CRISPR/Cas9 technology. However, target sequence modification by CRISPR/Cas9 has not been reported in the plant mitochondrial genome, mtDNA. In plants, a type of male sterility known as cytoplasmic male sterility (CMS) has been associated with certain mitochondrial genes, but few genes have been confirmed by direct mitochondrial gene-targeted modifications. Here, the CMS-associated gene (mtatp9) in tobacco was cleaved using mitoCRISPR/Cas9 with a mitochondrial localization signal. The male-sterile mutant, with aborted stamens, exhibited only 70% of the mtDNA copy number of the wild type and exhibited an altered percentage of heteroplasmic mtatp9 alleles; otherwise, the seed setting rate of the mutant flowers was zero. Transcriptomic analyses showed that glycolysis, tricarboxylic acid cycle metabolism and the oxidative phosphorylation pathway, which are all related to aerobic respiration, were inhibited in stamens of the male-sterile gene-edited mutant. In addition, overexpression of the synonymous mutations dsmtatp9 could restore fertility to the male-sterile mutant. Our results strongly suggest that mutation of mtatp9 causes CMS and that mitoCRISPR/Cas9 can be used to modify the mitochondrial genome of plants.

Novel Hina alleles created by genome editing increase grain hardness and reduce grain width in barley.

The hordoindolina genes (Hina and Hinb) are believed to play critical roles in barley (Hordeum vulgare L.) grain texture. In this study, we created novel alleles of the Hina gene using CRISPR/Cas9 (Clustered regularly inter spaced short palindromic repeat-associated protein, CRISPR-Cas) genome editing. Mutagenesis of single bases in these novel alleles led to loss of Hina protein function in edited lines. The grain hardness index of hina mutants was 95.5 on average, while that of the wild type was only 53.7, indicating successful conversion of soft barley into hard barley. Observation of cross-sectional grain structure using scanning electron microscopy revealed different adhesion levels between starch granules and protein matrix. Starch granules were loose and separated from the protein matrix in the wild type, but deeply trapped and tightly integrated with the protein matrix in hina02 mutants. In addition, the grain width and thousand-grain weight of the hina02 mutant were significantly lower than those of the wild type.

Up-regulating microRNA-138-5p enhances the protective role of dexmedetomidine on myocardial ischemia-reperfusion injury mice via down-regulating Ltb4r1.

Both microRNAs (miRs) and dexmedetomidine (Dex) have been verified to exert functional roles in myocardial ischemia-reperfusion injury (MI/RI). Given that, we concretely aim to discuss the effects of Dex and miR-138-5p on ventricular remodeling in mice affected by MI/RI via mediating leukotriene B4 receptor 1 (Ltb4r1). MI/RI mouse model was established by ligating left anterior descending coronary artery. The cardiac function, inflammatory factors and collagen fiber contents were detected after Dex/miR-138-5p/Ltb4r1 treatment. MiR-138-5p and Ltb4r1 expression in myocardial tissues were tested by RT-qPCR and western blot assay. The target relationship between miR-138-5p and Ltb4r1 was verified by online software prediction and luciferase activity assay. MiR-138-5p was down-regulated while Ltb4r1 was up-regulated in myocardial tissues of MI/RI mice. Dex improved cardiac function, alleviated myocardial damage, reduced inflammatory factor contents, collagen fibers, and Ltb4r1 expression while increased miR-138-5p expression in myocardial tissues of mice with MI/RI. Restored miR-138-5p and depleted Ltb4r1 improved cardiac function, abated inflammatory factor contents, myocardial damage, and content of collagen fibers in MI/RI mice. MiR-138-5p directly targeted Ltb4r1. The work evidence that Dex could ameliorate ventricular remodeling of MI/RI mice by up-regulating miR-138-3p and down-regulating Ltb4r1. Thus, Dex and miR-138-3p/Ltb4r1 may serve as potential targets for the ventricular remodeling of MI/RI.

Risk Factors for Mortality of Inpatients with Pseudomonas aeruginosa Bacteremia in China: Impact of Resistance Profile in the Mortality.

PURPOSE: Pseudomonas aeruginosa bacteremia presents a severe challenge to hospitalized patients. However, to date, the risk factors for mortality among inpatients with P. aeruginosa bacteremia in China remain unclear. PATIENTS AND METHODS: This retrospective multicenter study was performed to analyze 215 patients with culture-confirmed P. aeruginosa bacteremia in five healthcare centers in China during the years 2012-2019. RESULTS: Of 215 patients with P. aeruginosa bacteremia, 61 (28.4%) died during the study period. Logistic multivariable analysis revealed that cardiovascular disease (OR=3.978, P=0.001), blood transfusion (OR=5.855, P<0.001) and carbapenem-resistant P. aeruginosa (CRPA) phenotype (OR=4.485, P=0.038) constituted the independent risk factors of mortality. Furthermore, both CRPA and multidrug-resistant P. aeruginosa (MDRPA) phenotypes were found to be significantly associated with 5-day mortality (Log-rank, P<0.05). CONCLUSION: This study revealed a high mortality rate amongst hospitalized patients with P. aeruginosa bacteremia, and those with cardiovascular diseases, CRPA and MDRPA phenotypes, should be highlighted and given appropriate management in China.

Effect of Pregnane X Receptor*1B genetic polymorphisms on postoperative analgesia with fentanyl in Chinese patients undergoing gynecological surgery.

BACKGROUND: The purpose of the study was to investigate the effects of the pregnane X receptor (PXR)*1B polymorphisms on CYP3A4 enzyme activity and postoperative fentanyl consumption in Chinese patients undergoing gynecological surgery. METHODS: A total of 287 females of Han ethnicity, aged 20 to 50 years old, ASA I or II, scheduled to abdominal total hysterectomy or myomectomy under general anesthesia were enrolled. The analgesic model used was fentanyl consumption via patient-controlled intravenous analgesia (PCIA) in the post-operative period. Additionally, pain was assessed using a visual analog score (VAS). Pain scores, occurrence of adverse reactions and consumption of fentanyl were recorded during the 24 h postoperative period. The enzyme activity of CYP3A4 was evaluated by measuring the plasma ratio of 1'-hydroxymidazolam to midazolam 1 h after intravenous administration of 0.1 mg/kg midazolam. PXR genotyping was performed by direct DNA sequencing and the PXR * 1B haplotype was analyzed via PHASE V.2.1 software. RESULTS: The polymorphism frequency of PXR11156A > C/11193 T > C and 8055C > T were 49.6 and 49.3%, and the rate of PXR * 1B haplotype was 48.8% in our study. None of the pain scores, consumption of fentanyl 24 h post-operatively or enzyme activity of CYP3A4, showed differences among different genotypes. CONCLUSIONS: PXR11156A > C, PXR11193T > C, PXR8055C > T or the PXR * 1B haplotype do not appear to be important factors contributing to CYP3A4 activity and interindividual variations in postoperative fentanyl consumption in Han female patients undergoing gynecological surgery. TRIAL REGISTRATION: The DNA samples were obtained since 2007 to 2010 year in our hospital, there was no registration at that time. So this section is not applicable to our research.

High Prevalence and Properties of Enterotoxin-Producing Staphylococcus aureus ST5 Strains of Food Sources in China.

Staphylococcus aureus with the ability of staphylococcal enterotoxins (SEs) production is one of the most common causes of bacterial foodborne outbreaks worldwide. In our study, 336 S. aureus isolates were recovered from 3476 food samples during 2010-2014. A total of 86 S. aureus isolates were proved to be enterotoxin-producing strains with PCR and enzyme-linked immunosorbent assay. In the 86 isolates, 20 STs were identified using multilocus sequence typing (MLST) and 20 isolates were typed as sequence type 5 (ST5), which was the most prevalent ST using MLST. There were six SE profiles and high carrier rates of sec (50%) and sed (75%) genes in the 20 S. aureus ST5 isolates. Additionally, 8 antibiotic resistance patterns were observed, and 10 multidrug-resistant isolates (50%) and 4 methicillin-resistant S. aureus isolates were identified. Our findings illustrate high prevalence of S. aureus ST5 isolates from food sources and diversity in SE profiles and antibiotic resistance patterns. These results indicate that great difference in the ability of obtaining SE production and antimicrobial resistance may exist between different genetic lineages of S. aureus strains.

Effect of ABCB1 rs12720464 and rs1055302 polymorphisms in Chinese patients on the time course of action of rocuronium administered as a single dose.

OBJECTIVE: To determine whether ABCB1 gene polymorphisms affect the time course of action of rocuronium in Chinese patients. METHODS: This study included 105 unrelated Chinese patients undergoing general anesthesia with propofol, fentanyl, and rocuronium. Neuromuscular monitoring was performed with calibrated acceleromyography. Patients were allowed to recover spontaneously from the neuromuscular block. The time interval between the first maximum depression of the train of four (TOF) and spontaneous recovery TOF ratio of 0.25/0.7/0.8/0.9 was recorded. The Sequenom MassArray® single-nucleotide polymorphism (SNP) detection technology was used to detect the genotypes of the ABCB1 rs12720464, rs1055302. Demographic and non-genetic clinical data were also collected. RESULTS: In the present study, the mean time to spontaneous recovery of TOF ratio 0.8/0.9 in ABCB1 rs12720464 GG genotype was longer compared to that observed in ABCB1 rs12720464 AG genotype (56.77 ± 14.23 minutes vs. 49.50 ± 10.49 minutes, and 62.58 ± 18.16 minutes vs. 53.20 ± 12.56 minutes, respectively, p < 0.05). Further, the time to spontaneous recovery of TOF 0.7/0.8/0.9 in ABCB1 rs1055302 GG genotype was longer than that in ABCB1 rs1055302 AG genotype (52.00 ± 12.10 minutes vs. 44.83 ± 7.38 minutes, 55.96 ± 13.92 minutes vs. 46.83 ± 7.67 minutes, 61.66 ± 17.70 minutes vs. 49.50 ± 8.44 minutes, respectively, p < 0.05). CONCLUSION: In Chinese patients who were administered a single dose of rocuronium, the genetic variants ABCB1 rs12720464, and rs1055302 contribute to the individual< variability of time course of action.

Prevalence and Characteristics of Enterotoxin B-Producing Staphylococcus aureus Isolated from Food Sources: A Particular Cluster of ST188 Strains was Identified.

The aim of this study was to investigate the prevalence and characteristics of staphylococcal enterotoxin B (SEB) producing Staphylococcus aureus (S. aureus) isolated from food sources. A total of 412 S. aureus isolates were recovered from 1970 milk and dairy samples (n = 236) and 2450 meat samples (n = 176) in China from 2009 to 2014. Of the 412 isolates, 124 isolates were tested positive for 1 or more classical staphylococcal enterotoxin (SE) genes using PCR, and 31 isolates were positive for seb gene and further proved to be SEB-producing. Four SE profiles were observed among 31 SEB-producing isolates when investigated using ELISA kit, that is, SEB (16 isolates), SEA+SEB (6 isolates), SEB+SEC (6 isolates), and SEB+SED (3 isolates). Thirteen sequence types (STs) were identified in the 31 SEB-producing S. aureus isolates using multilocus sequence typing (MLST). The 3 most detected STs were ST1 (7 isolates), ST188 (6 isolates), ST59 (3 isolates). Two distinct clusters were identified by pulsed-field gel electrophoresis (PFGE), each of which showed excellent consistency with ST188 and ST1 achieved by MLST, respectively. In summary, this study reveals that various SE profiles are observed in SEB-producing S. aureus isolates and the great part of SEB-producing S. aureus isolates are showed as clusters. Especially, a particular cluster of ST188 strains was observed in SEB-producing S. aureus isolates which was associated with outbreaks of SFP and needs further attention.

Tracheal rupture related to endotracheal intubation after thyroid surgery: a case report and systematic review.

Tracheobronchial rupture is an uncommon but potentially serious complication of endotracheal intubation. The most likely cause of tracheal injury is massive overinflation of the endotracheal tube cuff and pre-existing tracheal wall weakness. We review the relevant literature and predisposing factors contributing to this complication. Only articles that reported at least the demographic data (age and sex), the treatment performed and the outcome were included. Papers that did not detail these variables were excluded. We also focus on a case of tracheal laceration after tracheal intubation in a patient with severe thyroid carcinoma. This patient received surgical repair and recovered uneventfully. Two hundred and eight studies that reported cases or case series were selected for analysis. Most of the reported cases (57·2%) showed an uneventful recovery after surgical therapy. The overall mortality was 19·2% (40 patients). Our patient too recovered without any serious complication. Careful prevention, early detection and proper treatment of the problem are necessary when tracheal rupture occurs. The morbidity and mortality associated with tracheal injury mandate a high level of suspicion and expedient management.

CYP3A4 * 1G Genetic Polymorphism Influences Metabolism of Fentanyl in Human Liver Microsomes in Chinese Patients.

PURPOSE: This study aimed to investigate whether CYP3A4*1G genetic polymorphism influences the metabolism of fentanyl in human liver microsomes in Chinese patients. METHODS: The human liver microsomes were obtained from 88 hepatobiliary surgery patients who accepted liver resection surgery in this study. A normal liver sample (confirmed by the Department of Pathology) was taken from the outer edge of the resected tissue. The metabolism of fentanyl in human liver microsomes was studied. The concentration of fentanyl was measured by high performance liquid chromatography. The CYP3A4*1G variant allele was genotyped using the PCR restriction fragment length polymorphism method. RESULTS: The frequency of the CYP3A4*1G variant allele was 0.188 in the 88 Chinese patients who had received hepatobiliary surgery. The metabolic rate of fentanyl in patients homozygous for the *1G/*1G variant (0.85 ± 0.37) was significantly lower than that in patients bearing the wild-type allele *1/*1 (1.89 ± 0.58) or in patients heterozygous for the *1/*1G variant (1.82 ± 0.65; p < 0.05). There were no gender-related differences in the metabolic rate of fentanyl (p > 0.05) nor was there any correlation between age and metabolic rate of fentanyl (p > 0.05). Results from different hepatobiliary diseases showed no significant difference in the metabolic rate of fentanyl (p > 0.05). The difference of CYP3A4 mRNA among different CYP3A4*1G variant alleles was significant (p < 0.05). There was positive correlation between CYP3A4 mRNA and metabolic rate of fentanyl (p < 0.01). CONCLUSIONS: CYP3A4*1G genetic polymorphism decreases the metabolism of fentanyl. There is a positive correlation between CYP3A4 mRNA level and metabolism of fentanyl.

Effects of low-flow sevoflurane anesthesia on renal function in low birth weight infants.

BACKGROUND: Low-flow sevoflurane anesthesia has been shown to influence renal function in rats, but not in adult humans. Presently, no study has assessed the effects of sevoflurane on renal function in low birth weight infants. Our aim was to study the renal function in low birth weight infants undergoing surgery with low-flow sevoflurane anesthesia. METHODS: Forty infants graded as American Society of Anesthesiologists (ASA) grade I or II undergoing abdominal surgery were selected. After the induction of anesthesia, they received sevoflurane semi-closed inhalation anesthesia with an oxygen flow rate of 1 L/minute. According to patient vital signs, in-tidal sevoflurane concentration was maintained at 2.5%-4.0%. Peripheral vein blood samples and urine specimens were obtained before surgery (T0), at the end of surgery (T1), and 24 (T2), 48 (T3), and 72 hours (T4) after surgery. Serum creatinine (Cr), blood urea nitrogen (BUN), urinary retinol binding protein (RBP), and ß-N-acetyl-glucosaminidase (NAG) levels were determined at these time points. Also, a temperature probe was inserted into the center of a soda lime canister and temperature readings were obtained. RESULTS: There were no significant differences in Cr and BUN before and after surgery (P > 0.05). However, RBP and NAG levels increased after surgery (P < 0.05), but returned to preoperative levels 72 hours (T4) after surgery. The highest soda lime temperature was 37.3 ± 3.1°C. CONCLUSIONS: Low-flow sevoflurane semi-closed inhalation anesthesia has no significant effect on the renal function of low birth weight infants.

Genome-wide identification and functional annotation of Plasmodium falciparum long noncoding RNAs from RNA-seq data.

The life cycle of Plasmodium falciparum is very complex, with an erythrocytic stage that involves the invasion of red blood cells and the survival and growth of the parasite within the host. Over the past several decades, numbers of studies have shown that proteins exported by P. falciparum to the surface of infected red blood cells play a critical role in recognition and interaction with host receptors and are thus essential for the completion of the life cycle of P. falciparum. However, little is known about long noncoding RNAs (lncRNAs). In this study, we designed a computational pipeline to identify new lncRNAs of P. falciparum from published RNA-seq data and analyzed their sequences and expression features. As a result, 164 novel lncRNAs were found. The sequences and expression features of P. falciparum lncRNAs were similar to those of humans and mice: there was a lack of sequence conservation, low expression levels, and high expression coefficient of variance and co-expression with nearby coding sequences in the genome. Next, a coding/noncoding gene co-expression network for P. falciparum was constructed to further annotate the functions of novel and known lncRNAs. In total, the functions of 69 lncRNAs, including 44 novel lncRNAs, were annotated. The main functions of the lncRNAs included metabolic processes, biosynthetic processes, regulation of biological processes, establishment of localization, catabolic processes, cellular component organization, and interspecies interactions between organisms. Our results will provide clues to further the investigation of interactions between human hosts and parasites and the mechanisms of P. falciparum infection.

Influence of CYP3A5*3 polymorphism and interaction between CYP3A5*3 and CYP3A4*1G polymorphisms on post-operative fentanyl analgesia in Chinese patients undergoing gynaecological surgery.

BACKGROUND AND OBJECTIVE: Fentanyl is metabolised by cytochrome P450 (CYP) 3A4 and CYP3A5. Our previous work demonstrated that the CYP3A4*1G polymorphism significantly affects the post-operative fentanyl analgesic effect in Chinese women undergoing gynaecological surgery. However, whether CYP3A5*3, a frequent single nucleotide polymorphism of CYP3A5 in Chinese people, affects the post-operative analgesic effect of fentanyl is unclear. In this study, we assessed the influence of the CYP3A5*3 polymorphism and the interaction of the CYP3A5*3 and CYP3A4*1G polymorphisms on post-operative fentanyl analgesia in Chinese women undergoing gynaecological surgery. METHODS: We enrolled 203 women scheduled for abdominal total hysterectomy or myomectomy under general anaesthesia. Intravenous fentanyl patient-controlled analgesia was provided post-operatively for adequate analgesia. Pain scores and fentanyl consumption were recorded 24 h post-operatively. Midazolam was used as a probe drug, and CYP3A activity was measured by plasma ratio of 1'-hydroxymidazolam to midazolam 1 h after intravenous administration of 0.1 mg kg-1 midazolam. Blood samples were genotyped for the CYP3A5*3 polymorphism. RESULTS: The frequency of the CYP3A5*3 allele was 72.4% in 203 patients. CYP3A activity did not differ among CYP3A5*3 genotypes. Fentanyl consumption 24 h post-operatively was lower with CYP3A5*1/*3 and CYP3A5*3/*3 polymorphisms than with CYP3A5*1/*1, but the differences were not statistically significant. However, combined with CYP3A4*1G polymorphism, post-operative fentanyl consumption at 24 h was significantly lower for the CYP3A5*1/*3 or CYP3A5*3/*3 group than the CYP3A5*1/*1 group. CONCLUSION: CYP3A5*3 is not the main genetic factor contributing to interindividual variation in the post-operative analgesic effect of fentanyl in Chinese women undergoing gynaecological surgery; an interaction between CYP3A5*3 and CYP3A4*1G polymorphisms can significantly influence the post-operative effect.

The wild-type hepatitis C virus core inhibits initiation of antigen-specific T- and B-cell immune responses in BALB/c mice.

In this study, the effects of wild-type and deletion mutant hepatitis C virus (HCV) core proteins on the induction of immune responses in BALB/c mice were assessed. p2HA-C145-S23, encoding a core protein with the C-terminal 46 amino acids truncated, significantly produced stronger antibody and cellular responses than p2HA-C191-S23. The induction of immune responses by p2HA-C145-S23 was dose dependent. However, increasing the doses or repeated administration did not enhance immune responses by the wild-type core protein. In addition, p2HA-C191-S23 was apparently able to interfere with the priming of specific immune responses by p2HA-C145-S23 when the two were coadministered. These results demonstrated that the wild-type HCV core protein itself could inhibit the priming of immune responses in the course of a DNA vaccination, whereas the truncated HCV core protein could provide potential applications for the development of DNA- and peptide-based HCV vaccines.

CYP3A4*1G genetic polymorphism influences CYP3A activity and response to fentanyl in Chinese gynecologic patients.

PURPOSE: To investigate whether the CYP3A4*1G genetic polymorphism contributes to the variability in CYP3A activity and response to fentanyl. METHODS: One hundred and forty-three gynecologic patients who were scheduled to undergo abdominal total hysterectomy or myomectomy with general anesthesia were enrolled in this study. Intravenous fentanyl patient-controlled analgesia was provided postoperatively for satisfactory analgesia. The degrees of pain at rest during PCA treatment were assessed with visual analog scale. The fentanyl consumption and occurrence of any adverse effects were recorded in the first 24 h postoperatively. CYP3A activity was measured by plasma 1'-hydroxymidazolam-to-midazolam ratio 1 h after intravenous administration of 0.1 mg/kg midazolam. CYP3A4*1G variant allele was genotyped using the polymerase chain reaction-restriction fragment length polymorphism method. RESULTS: The frequency of the CYP3A4*1G variant allele was 0.269 in 143 Chinese gynecologic patients. The activity of CYP3A4 in patients homozygous for the *1G/*1G variant (0.34 +/- 0.15) was significantly lower than that in patients bearing the wild-type allele (*1/*1) (0.46 +/- 0.14) or in patients heterozygous for the *1/*1G variant (0.46 +/- 0.12) (P < 0.05). The patients with the CYP3A4*1G/*1G genotype needed less fentanyl (227.8 +/- 55.2 microg) to achieve pain control than patients carrying the CYP3A4*1/*1 (381.6 +/- 163.6 microg) and CYP3A4*1/*1G (371.9 +/- 180.1 microg) genotypes (P < 0.05) during the first 24 h postoperatively. There was no significant difference in incidence of adverse events among the different genotype groups (P > 0.05). CONCLUSIONS: CYP3A4*1G genetic polymorphism decreases CYP3A activity and fentanyl consumption for postoperative pain control.

Relationship between perioperative inflammatory response and postoperative cognitive dysfunction in the elderly.

After surgery and anesthesia, many elderly patients show a decline in cognitive function. This condition is called postoperative cognitive dysfunction (POCD). POCD, a distressing complication after surgery, is independently associated with poor short-term and long-term outcomes. Many pathophysiological mechanisms have been implicated in development of POCD, but the exact cascade leading to its development is unclear. Animal studies show that cytokines from inflammatory response are involved in with cognitive dysfunction. Simultaneously, emerging evidences indicate that inflammatory response represents a potential pathogenic factor in many central cognitive diseases. A similar story may be occurring during perioperative process in patients. Surgical trauma, anesthesia, and stress response induced perioperative nonspecific inflammatory response. We hypothesize that perioperative inflammatory response promotes the development of POCD in elderly surgical patients, and some measures against perioperative inflammatory response should be considered as a new pathway to prevention of POCD.

[Study of the effect of hepatitis C virus core protein on interferon-induced antiviral genes expression and its mechanisms].

To study the effect of HCV core protein on the interferon-induced antiviral genes expression and its mechanisms. Methods HepG2 cells were transiently transfected with HCV core protein expression plasmid and the blank plasmid respectively. RT-PCR was used to analyze the effect of HCV core protein on PKR and 2'-5'OAS expression. The effect of HCV core protein on ISRE-medicated gene expression was detected by luciferase activity assay. Western-blot assay was performed to observe the change of mRNA and protein levels of SOCS3, STAT1 and p-STAT1 following HCV core expression. In the presence of HCV core protein, the transcription of PKR and 2'-5' OAS are down-regulated. ISRE-medicated reporter gene expression and STAT1 phosphorylation were inhibited. The transcription and expression of SOCS3 were induced compared with blank plasmid-transfected group. In HepG2 cells, HCV core protein can down-regulate the expression of some interferon-induced antiviral genes, which involves the induction of SOCS3 and the inhibition of STAT1 phosphorylation.