The CRISPR/Cas9 based prime editing (PE) technique enables all 12 types of base substitutions and precise small DNA deletions or insertions without generating DNA double-strand breaks. Prime editing has been successfully applied in plants and plays important roles in plant precision breeding. Although plant prime editing (PPE) can substantially expand the scope and capabilities of precise genome editing in plants, its editing efficiency still needs to be further improved. Here, we review the development of PPE technique, and introduce structural composition, advantages and limitations of PPE. Strategies to improve the PPE editing efficiency, including the T_m-directed PBS length design, the RT template length, the dual-pegRNA strategy, the PlantPegDesigner website, and the strategies for optimizing the target proteins of PPE, were highlighted. Finally, the prospects of future development and application of PPE were discussed.
CRISPR-Cas Systems/genetics* ; DNA ; Gene Editing ; Genome, Plant/genetics* ; Plant Breeding ; Plants/genetics*

Tree shrew is a novel and high-quality experimental animal model. In this study, the real-time polymerase chain reaction methods were established to detect infection-related cytokines interleukin-6 (IL-6), IL-8, IL-10, IL-17A, interferon-γ (IFN-γ) and housekeeping gene glyceraldehyde-phosphate dehydrogenase ( ) of tree shrew. The results indicated that the establised methods had good specificity. The high point of the linear range of these reagents reached 1 × 10 copies, and the low points ranged from 10 copies (IL-6, IL-17A), 100 copies (IL-10, ) to 1 000 copies (IL-8, IFN-γ). In this interval, the linear correlation coefficient of each reagent was greater than 0.99. The lowest detectable values of IL-6, IL-8, IL-10, IL-17A, IFN-γ and were 8, 8, 4, 8, 128 and 4 copies, respectively. The results showed that the established detection methods had good specificity, sensitivity and wide linear range. The methods were suitable for detection of multiple concentration range samples, and could be used for the subsequent studies of tree shrew cytokines.
Animals ; Cytokines ; analysis ; Real-Time Polymerase Chain Reaction ; Shrews

Objective To explore the role of cell apoptosis pathway in alcoholic pancreatitis.Methods C57BL/J mice were divided into control group (NC) and Alcohol group (AC),Acute pancreatitis group (AP) and Alcoholic acute pancreatitis group(AAP).Alcohol treatment was 10％ w/v ethanol feeding for 2 d,15％ w/v ethanol for 5 d,and then 20％ w/v ethanol until 13 weeks.AP model was established by the intraperitoneal injection of 50μg caerulein/kg body weight once an hour for a total of 7 times.Blood samples were collected for detecting serum amylase and lipase activity.Part pancreatic tissue was collected and the wet and dry weight were both measured to calculate the water content.The routine pathological exanination of the pancreatic tissues were conducted.The expression of apoptosis associated protein caspase3 and caspase8 was determined by Western blot.And cell apoptosis was determined using TUNNEL method.Results The level of serum amylase in NC group,AC group,AP group and AAP group were(3 630 ± 259),(3 196 ± 187),(35 955 ± 4607) and (53 607 ± 3 848) U/L;the level of serum lipase were (502 ± 41),(745 ± 42),(7 346 ± 665) and(12 764 ± 2 544) U/L;the water content were (70.2 ± 3.1) ％,(69.6 ± 2.0) ％,(78.2 ± 1.5) ％ and(85.0 ± 3.0) ％ and (12.75 ± 0.25);the expression of caspase3 were (1.017 ± 0.0784),(1.287 ± 0.097),(178 ± 0.07785) and (0.2443 ± 0.0243);the expression of caspase8 were (0.8289 ± 0.0096),(0.5985 ±0.0735),(1.27 ±0.08) and (0.145 ±0.015);the number of apoptotic cells were 1,6,214,97/10 high power field.The pathological score of pancreas injure in NC group,AC group,AP group and AAP group were 0,0,(7 ± 0.4) and (12.8 ± 0.3),respectively.Serum anylase,lipase,water content and pathological scores in AP group were obviously higher than those in NC group (P ＜ 0.05),which in AAP group were also obviously higher than those in AP group,and all the differences were statistically significant (all P ＜0.05).Compared with NC group,the expressions of apoptosis associated protein caspase3 and caspase8 and the number of apoptotic cells were obviously increased in AP group,which were obviously higher than those in AAP group,but the expression of caspase3 and caspase8 in AAP group were decreased compared with NC group,and all the differences were statistically significant (all P ＜ 0.05).Conclusions Chronic alcohol exposure may aggravate the severity of pancreatitis,and the inhibition of apoptosis pathway and the enhancement of acinar cell necrosis may be involved in this process.

Objective:To collect the clinical materials of primary liver cancer patients treated by gamma knife and analyze the treatment methods as well as short-term efficacy,and to provide a reference for the clinical treatment of primary liver cancer patients.Methods:A total of 633 patients with inoperable primary liver cancer were treated by body gamma knife.According to the TNM staging method of Union for International Cancer Control(UICC), there were 351 cases with clear TNM staging.Among them, there were 251 cases (71.5%) at T3 stage and 57 cases(16.2%) at T4 stage.The prescription dose of 200-600 cGy each time to the 40%-85% dose line covering the planned target volume(PTV), this program was performed 5 times per week, and the number of treatment ranged from 2 to 13.The biochemical and imaging changes were observed 2-3 months after treatment to evaluate the short-term efficacy.Results:During the treatment, 229(36.2%)patients had adverse reactions,100 (15.8%) patients appeared the reduced white blood cells, and 137 (21.6%) patients appeared the reduced platelets.On discharge from the hospital, 601 patients were improved, 22 patients had no obvious change, 5 cases were worse, and 5 cases died.The proportion of improved patients who received the cumulative dose between 3 000 cGy to 4 000 cGy was higher than those who received the cumulative dose less than 3 000 cGy(P<0.05).After treatment, the curative effects were evaluated in 45 patients during two and three months, including PR 77.8%(35/45) and SD 22.2%(10/45), and the total effective rate was 77.8%(35/45).No statistical differences in effective rates were found between different tumor diameter, single dose, and cumulative dose groups(P>0.05).Conclusion:The proportion of adverse reactions in the primary liver cancer patients treated with body gamma knife is relatively low and the short-term efficacy is ideal.Body gamma knife treatment is a safe and effective treatment method for the primary liver cancer patients.

Objective To evaluate the clinical efficacy and safety of stereotactic radiotherapy (SRT) combined with bevacizumab for brain metastases in patients with lung adenocarcinoma.MethodsThe clinical data of 95 patients with brain metastases of lung adenocarcinoma were retrospectively analyzed, including 36 patients treated with SRT and bevacizumab (bevacizumab group) and 59 patients treated with SRT, corticosteroids, and mannitol (traditional drug group).The tumor response rate, peritumoral edema control rate, improvement in Karnofsky Performance Scale (KPS) score, and adverse reactions were analyzed.Results Compared with the traditional drug group, the bevacizumab group had significantly higher tumor response rate (P=0.033) and peritumoral edema control rate (P=0.000) at 1-4 weeks after treatment, as well as an obvious improvement in KPS score and reduced doses of corticosteroids and mannitol.In addition, the adverse reactions in the bevacizumab group were mild and controllable.Conclusions SRT combined with bevacizumab for brain metastases in patients with lung adenocarcinoma can achieve higher short-term tumor response rate and peritumoral edema control rate and improve patients' quality of life.

OBJECTIVETo investigate the changes of blood glucose in pancreatic cancer patients associated with diabetes mellitus (DM) after radiotherapy and analyze the related factors.

METHODSClinical data of 69 cases of stage I to III pancreatic carcinoma associated with DM were retrospectively analyzed. All patients were divided into four groups according to the history of DM, and the clinical characteristics and blood glucose changes after stereotactic radiotherapy were analyzed. The correlation between blood glucose and serum tumor markers was analyzed.

RESULTSNo significant differences were found in clinical characteristics of the four groups. The history of DM <12 months accounted for 39.1% of the pancreatic cancer patients, much more higher than that of patients with a longer disease course. After radiotherapy, the blood glucose changes were not significantly different between the two groups with DM history <12 months and 12-24 months (P = 0.519), but there were significant differences between the patients with DM history <12 months and those with DM history 25-36 months and >36 months (P < 0.05 for both). After radiotherapy, CA199/CEA was reduced and blood glucose was also improved in the patients with DM history <12 months and 12-24 months, and the reduction of CA199/CEA showed a significant positive correlation with the improvement of blood glucose (r = 0.834 and r = 0.660, P < 0.01 for both), however, no significant correlation was found between the two parameters in patients with DM history 25-36 months and >36 months (r = 0.319 and r = 0.439, P > 0.05 for both).

CONCLUSIONSHyperglycemia in diabetic patients with a disease course <24 months might be a clinical manifestation secondarily developed in pancreatic cancer patients. Therefore, patients with new onset diabetes should be closely followed-up for early detection of pancreatic cancer.

Aged ; Biomarkers, Tumor ; blood ; Blood Glucose ; Diabetes Mellitus ; blood ; Humans ; Male ; Middle Aged ; Pancreatic Neoplasms ; blood ; Retrospective Studies ; Time Factors

OBJECTIVESince 1986, the reference of birth weight for gestational age has not been updated. The aim of this study was to set up Chinese neonatal network to investigate the current situation of birth weight in China, especially preterm birth weight, to develop the new reference for birth weight for gestational age and birth weight curve.

METHODA nationwide neonatology network was established in China. This survey was carried out in 63 hospitals of 23 provinces, municipalities and autonomous regions. We continuously collected the information of live births in participating hospitals during the study period of 2011-2014. Data describing birth weight and gestational age were collected prospectively. Newborn's birth weight was measured by electronic scale within 2 hours after birth when baby was undressed. The evaluation of gestational age was based on the combination of mother's last menstrual period, ultrasound in first trimester and gestational age estimation by gestational age scoring system.

STATISTICAL ANALYSISthe growth curve was drawn by using LMSP method, which was conducted in GAMLSS 1.9-4 software package in R software 2.11.1.

RESULTA total of 159 334 newborn infants were enrolled in this study. There were 84 447 male and 74 907 female. The mean birth weight was (3 232 ± 555) g, the mean birth weight of male newborn was (3 271 ± 576) g, the mean weight of female newborn was (3 188 ± 528) g. The test of the variables' distribution suggested that the distribution of gestational age and birth weight did not fit the normal distribution, the optimal distribution for them was BCT distribution. The Q-Q plot test and worm plot test suggested that this curve fitted the distribution optimally. The male and female neonatal birth weight curve was developed using the same method.

CONCLUSIONUsing GAMLSS method to establish nationwide neonatal birth weight curve, and the first time to update the birth weight reference in recent 28 years.

Birth Weight ; China ; Female ; Gestational Age ; Humans ; Infant, Low Birth Weight ; Infant, Newborn ; Male

Objective To evaluate the preliminary clinical efficacy of bevacizumab for cerebral radiation necrosis (CRN). Methods Nineteen patients with CRN for whom the treatment with steroids and mannitol failed were retrospectively analyzed with a total of 22 lesions. Except for 5 lesions confirmed by pathological evidence, all lesions were confirmed by the following imaging evidence:1. computed tomography (CT)?or magnetic resonance imaging (MRI)?enhanced lesions showed loss of tension and were accompanied by substantial edema;2. CT?or MRI?enhanced lesions had a low perfusion pressure;3. magnetic resonance spectroscopy indicated that the enhanced areas had a decreased choline peak; 4. positron emission tomography showed that the fluorodeoxyglucose uptake was substantially reduced in the enhanced areas. All patients were given 5 mg/ kg bevacizumab at an interval of 14 days for 2?6 cycles. MRI examination was performed in each cycle before treatment, and the enhanced lesions on T1?weighted images ( T1 WI) and edema on T2?weighted images (T2 WI) were compared before and after treatment. The clinical symptoms, Karnofsky Performance Status ( KPS), and adverse reactions in all patients were evaluated. Comparison before and after treatment was performed by paired t test. Results All 19 patients completed the treatment successfully and there were no severe adverse reactions. The clinical symptoms of patients were substantially improved after the second cycle of treatment, and the KPS score increased by 26?? 8 on average. The visible volume of enhanced lesions on MRI T1 WI was significantly reduced by 54?? 8% after treatment (P= 0?? 000), while the visible volume of edema on MRI T2 WI was reduced by 80?? 7% after treatment (P= 0?? 000). The follow?up time ranged from 3 to 12 months with a mean value of 5?? 6 months. Eleven patients kept clinical improvement in CRN, four patients had recurrence, and four patients died from tumor progression. Conclusions Bevacizumab is preliminarily confirmed to substantially improve the clinical symptoms and quality of life in patients with CRN.

Objective To explore the different probability of carcinoma and the necessity of treatment in cervical columnar ectopy (CCE),normal smooth cervix and cervical intraepithelial neoplasia(CIN) by investigating the different expression of matrix metalloproteinase-9(MMP-9).Methods The expression of MMP-9 was detected by using immunohistochemical method in 30 cases of CCE,28 cases of CIN and 30 cases of normal smooth cervix from 2007 to 2012.Results The positive expression of MMP-9protein in CIN was 96.4％ (27/28),CCE was 23.3％ (7/30) and normal smooth cervix was 20.0％ (6/30)respectively.The positive expression of MMP-9 protein was higher in CIN than CCE and normal smooth cervix,the difference has statistical significance (P＜ 0.01),there was no significant difference between CCE and normal smooth cervix (P ＞ 0.05).Conclusions CCE,which was a physiological performance of cervical erosion,should not be overtreated.But the women of childbearing age should be routinely performed in cervical cancer screening.

This study was aimed to examine the expression of apoptosis-associated gene Fas in HeLa cell, explore the effects of the co-immobilized cytokines (tumor necrosis factor-alpha and interferon-gamma), and probe the potential mechanism of action. The preparation and application of the research couple IFN-gamma and TNF-alpha to the polystyrene cell culture plate were performed using the Photo-immobilization method, with different doses (20 ng/well and 200 ng/well) and synthesized optical active material. HeLa cells were treated with cytokines for two dose and 1, 3, 6 days. The result showed that the free cytokines induced HeLa apoptosis quickly, yet the HeLa apoptosis induced by co-immobilized cytokines had longer effect.
Apoptosis ; drug effects ; genetics ; Drug Synergism ; HeLa Cells ; Humans ; Immobilized Proteins ; chemistry ; pharmacology ; Interferon-gamma ; chemistry ; pharmacology ; Tumor Necrosis Factor-alpha ; chemistry ; pharmacology ; Up-Regulation ; fas Receptor ; metabolism