Group Ⅱ introns are self-splicing ribozymes, which insert directly into target sites in DNA with high frequency through "retrohoming". They specifically and efficiently recognize and splice DNA target sites, endowing themselves with great potential in genetic engineering. This paper reviewed the gene targeting principle of group Ⅱ introns and the application in microbial genetic modification, and then analyzed the limitations of them in multi-functional gene editing and eukaryotes based on the "retrohoming" characteristics and the dependence on high Mg²⁺ concentration. Finally, we dissected the potential of group Ⅱ introns in the development of novel gene editing tools based on our previous research outcome and the structural characteristics of the introns, hoping to provide a reference for the application of group Ⅱ introns in biotechnology.
DNA ; Eukaryota ; Gene Targeting ; Introns/genetics* ; RNA, Catalytic/genetics*

Objective:To explore the differences in intravascular ultrasound results in elderly coronary heart disease(CHD)patients with different uric acid levels.Methods:A total of 145 elderly patients diagnosed with CHD in our hospital from December 2017 to May 2020 were included as study subjects.Uric acid levels were measured and intravascular ultrasound examination was conducted in all patients.They were divided into different groups based on uric acid levels: Group A(uric acid≤199 μmol/L), Group B(uric acid 200~399 μmol/L)and Group C(uric acid≥400 μmol/L). Data from intravascular ultrasound-derived indexes were analyzed and compared between the three groups.Results:There was no significant difference in the degree of left main stenosis between Group A and Group B, but it was less severe in both groups than in Group C( F=5.625, P=0.039). Plaque fibrous cap thickness showed no significant difference between Group B and Group C, but it was smaller than in Group A( F=7.825, P=0.020). Group C had the largest plaque area and maximum thickness among the three groups, followed by Group B[(11.12±1.73)mm 2 and(1.76±0.24)mm]and Group A[(8.29±3.14)mm 2 and(1.38±0.09)mm]( F=6.384 and 6.827, P=0.028 and 0.015). Conclusions:Elevated uric acid levels in elderly CHD patients can increase the area and thickness of plaques, and reduce plaque fibrous cap thickness, leading to an increased risk of formation of unstable plaques, which can be life-threatening for these patients.Thus, monitoring and managing uric acid levels should be stressed in elderly CHD patients.

Objective: To investigate the action and antioxidant effects of CB2 agonist AM-1241 on rat hepatic stellate cell line (HSC-T6). Methods: HSC-T6 was randomly divided into four groups: control group, oxidative stress group, AM-1241 intervention group and AM-1241+AM-630 antagonist group. Survival rate of HSC-T6 was detected by thiazolyl blue assay under 24 h interventions with 0, 20, 50, 80 μmol/L AM-1241 and 0, 10, 20, 30, 40 μmol/L AM-630, respectively. Besides control group, the remaining groups were well cultured in low-glucose DMEM containing 100 mU/L glucose oxidase (GO) for 12 h to prepare the oxidative stress model. Then, AM-1241 intervention group was treated with 50 μmol/L low-glucose DMEM medium. After incubation for 12 h, the AM-1241+AM-630 antagonist group was treated with CB2 antagonist AM-630 (20 μmol/L) for 2 h, and cultured with 50 μmol/L AM-1241 in complete low-glucose medium for 12 h. The optimal drug concentration was selected according to the cell viability considered by the experiment results. Type III collagen (C III) content in the HSC-T6 supernatant was detected by enzyme-linked immunosorbent assay. Glutathione (GSH) content in HSC-T6 was detected by spectrophotometry. CB2 and heme oxygenase-1(HO-1) in each group of HSC-T6 were detected by western blotting. Results: HSC-T6 proliferation was inhibited in each group of AM-1241 in a concentration-dependent manner (P < 0.05). The inhibition was highest at 80μmol/L, and the cell survival rate was (41.61% ± 3.13%) (P < 0.05). AM-630 concentration group had no significant inhibitory effect on the proliferation of HSC-T6 (P > 0.05). HSC-T6 expressed CB2 receptor in each group. The expression level of CB2 in the AM-1241 intervention group was higher compare with control group (P < 0.05).The expression of Col III were significantly higher in oxidative stress group (P < 0.05) than in control group, and the expression of Col III of AM-1241 intervention group was significantly lower than that in oxidative stress group (P < 0.05). Col III level in AM-1241+AM-630 antagonistic group was significantly higher than that in AM-1241 intervention group (P < 0.05). There was no significant difference between AM-1241+AM-630 antagonistic group and oxidative stress group (P > 0.05). The content of GSH and HO-1 in oxidative stress group was higher (P < 0.05) than control group. The content of GSH and HO-1 in the AM-1241 intervention group was higher compared with oxidative stress group, while content of AM-1241 + AM-630 antagonist group was lower compared to AM-1241 intervention group (P < 0.05), and the differences were not statistically significant for oxidative stress group. Conclusion CB2 agonist AM-1241 can inhibit the proliferation and activation of HSC-T6 and its mechanism may activate the nuclear translocation of Nrf2 binding to HSC-T6, initiating the up-regulation of antioxidant enzymes HO-1 and GSH protein expression, and thus increase the antioxidant effect of HSC-T6.

Objective To investigate the feasibility of subtraction coronary computed tomography angiography (Sub-CCTA) for the diagnosis of coronary heart disease in the segment with severe calcification.Methods A retrospective analysis was performed on 27 patients who underwent clinically indicated digital subtraction angiography (DSA) and CCTA using a 320-detector row CT.Compared with the results of DSA,sensitivity,specificity,positive predictive value,negative predictive value and accuracy of Con-CCTA and Sub-CCTA were calculated.The clinical diagnostic accuracy of the two imaging methods was evaluated using the receiver operating characteristic (ROC) curve.The stenosis of coronary segments was divided into four grades (Ⅰ,Ⅱ,Ⅲ,Ⅳ).Kappa coefficient was used to measure agreement between two imaging methods.Image quality of 4-scale grade scoring method was used and t test was conducted.Results A total of 52 segments with severe calcification were evaluated.The scores of image quality in Con-CCTA and Sub-CCTA were 2.8 ± 0.5 and 3.4 ± 0.7,respectively.There was significant difference between them (t =5.9,P ＜ 0.05).Compared with the result of DSA as the golden standard,the Kappa coefficients were 0.55 and 0.81 respectively in Con-CCTA and Sub-CCTA for the quantitative evaluation of the severe calcified segments.The sensitivity,specificity,positive predictive value and negative predictive value and accuracy of Con-CCTA were 81.0％,63.1％,63.1％,81.1％ and 70.8 ％;and for Sub-CCTA they were 90.5 ％,85.2％,82.1 ％,92.0％ and 87.5 ％ respectively.Compared with Con-CCTA,the area under the ROC curve of Con-CCTA and Sub-CCTA were 0.84 (95％CI:0.70-0.93) and 0.96 (95％ CI:0.86-1.00),respectively,and the difference was statistically significant (P =0.03).Conclusions Sub-CCTA can improve the diagnostic accuracy of coronary artery stenosis in severe calcified segment.Application of subtraction technique in CCTA can reduce or even eliminate the artifacts caused by severe calcified plaque,and has a good clinical application prospect.

Objective: To investigate the clinical avalue of real time threshold manual triggering technique in the lower extremity arterial CT angiography .Methods :A total of 70 patients with clinically suspected lower extremity arterial disease underwent lower extremity arterial CT examination ,and the patients were randomly divided into two groups .In the first group 35 patients ,the automatic threshold triggering technique was used .After the threshold reached to 150 Hu ,6 s was delayed before scanning .The other group of 35 patients with manual triggering was used to observe the time density curve .When the peak value was stable ,scanning was started at once .The delay time of the two groups was analyzed and the image quality of the two groups was evaluated .The image quality evaluation included the branch level ,the smooth degree of the arterial edge , the interference of the vein and the CT value of the arterial segment .Results:There were significant differences in the starting delay time of the two trigger modes (P=0 .002 ,P<0 .05) ,and the auto trigger group was earlier than the manual triggering group .Between the two trigger modes ,there was no significant difference in the distal branches and vessel edge of abdominal‐iliac artery and femoral popliteal artery display ability .In the lower limb artery ,the image quality of manual triggering group was higher than that of the automatic trigger group ,the two groups had statistical significance (P=0 .004 , P=0 .003) .The same as to the distal branch and the edge of the vessel of the foot (P=0 .003 ,P=0 .003) .Two sets of images in the lower limb vein interference score had no significant difference ( P> 0 .05 ) .CT value had no significant difference in abdominal‐iliac artery ,femoral‐calf and popliteal arteries between the two groups ,while that in the dorsalis pedis artery had statistical significance between the two groups (P=0 .006) .Four levels of CT values in manual triggering group were higher than those in the automatic trigger group .Conclusions :Real time threshold manual triggering technique takes into account the individual differences of patients ,so the patient's delay scan time can be evaluated better and higher quality of lower extremity arterial images can be acquired .

Objective To investigate changes in expressions of activation antigens CD69 and HLA-DR in CD3+T lymphocytes in peripheral blood and skin lesions in patients with psoriasis vulgaris. Methods Peripheral blood samples were obtained from 20 patients with psoriasis vulgaris and 20 healthy controls, and skin specimens from the lesions of 15 out of the 20 patients and 10 healthy controls. Flow cytometry was performed to quantify the expressions of CD69 and HLA-DR in peripheral blood CD3+T cells, and an immunohistochemical study to measure the expression of HLA-DR in skin specimens. Statistical analysis was carried out by a two-sample t-test and Pearson correlation analysis with the SPSS 19.0 software. Results Compared with the healthy controls, the patients with psoriasis vulgaris showed increased expression rates of CD69 (4.70%± 1.90%vs. 1.56%± 0.95%, t=6.629, P<0.01)and HLA-DR (8.97%± 1.79% vs. 3.02% ± 1.15%, t= 6.204, P< 0.01)in peripheral blood. Pearson correlation analysis revealed that the percentage of CD3+HLA-DR+cells in peripheral blood was positively correlated with the psoriasis area and severity index (PASI)score (r=0.5626, P<0.05). The expression rate of HLA-DR was significantly higher in the dermis (64.87%± 17.31%vs. 19.80%± 5.69%, t=7.916, P<0.01), but lower in the epidermis(11.80%± 5.55%vs. 27.40%± 8.61%, t=5.479, P<0.01)in the psoriatic specimens compared with the control specimens. Immunohistochemically, HLA-DR was widely expressed in the dermis of psoriatic lesions, but mainly distributed around blood vessels in the control skin. Conclusions There is an aberrant activation of CD3+T cells in peripheral blood and inflammatory cells in skin lesions in patients with psoriasis vulgaris, and the percentage of CD3 +HLA-DR+ cells in peripheral blood is correlated with the severity of psoriasis vulagaris.

Objective:To investigate the correlations of Lauren classification and world health organization (WHO) classification of gastric cancer (GC) with microvascular density (MVD), matrix metalloproteinase-2 (MMP-2), matrix metalloproteinase-9 (MMP-9), vascular endothelial growth factor (VEGF), vascular endothelial growth factor receptor 1 (VEGFR1), vascular endothelial growth factor receptor 2 (VEGFR2), and p53. Methods:The clinical data of 89 patients with GC were collected. The collected specimens were categorized on the basis of Lauren classification and WHO classification. CD34/periodic acid-Schiff (PAS) double staining was performed to validate MVD. Immunohistochemistry was conducted to investigate the expression levels of MMP-2, MMP-9, VEGF, VEGFR1, VEGFR2, and p53. Results:MVD was not correlated with Lauren classification or WHO classification (P>0.05). Lauren typing was associated with the expression levels of MMP-9, VEGFR1, and p53 (P<0.05). WHO classification was not related to any of the factors (P>0.05). Cox proportional hazards model revealed that Lauren classification and WHO classification were the prognostic factors of overall survival (P<0.05). Conclusion:This research on tumor related factors, angiogenesis, and different classifications of GC may provide new methods to treat this disease.

Objective To evaluate the efficacy of narrow-band ultraviolet B (NB-UVB) for the treatment of early-stage mycosis fungoides (MF).Methods A retrospective study was performed on 98 patients with early-stage MF treated with NB-UVB from January 2003 to December 2013.The rate of and time to recurrence of MF were assessed,and long-term side effects were evaluated in these patients.Results Among 52 patients achieving complete remission,the median number of sessions of NB-UVB radiation was 30 (range,18-87),the median cumulative radiation dose was 24.16 J/cm2 (range,9.11-108.86 J/cm2),and the median follow-up duration was 41 months (range,11-90 months).Relapse occurred in 14 (26.9％) patients with the median time to recurrence being 8.2 months (range,3-60 months).The patients suffering from relapse achieved complete remission after treatment with NB-UVB again.Adverse reactions mainly included skin erythema and itching at the early stage of radiation,as well as temporary pigmentation after radiation.Cutaneous carcinoma occurred in none of these patients during 11-90 months of follow-up.Conclusions NB-UVB phototherapy appeared to be effective for the treatment of early-stage MF,and no cutaneous carcinoma occurred in these patients during the follow-up.Although MF may relapse in some cases,repeated NB-UVB phototherapy is still effective.

Objective:To compare the value of respiratory-triggered diffusion-weighted imaging with fat suppression(RTIR-DWI)and common breath-hold DWI(BH-DWI)for the differential diagnosis of pancreatic cancer and mass-like pancreatitis. Methods:A total of 30 cases of pancreatic cancer and 17 cases of mass-like pancreatitis,which were confirmed by surgical pa-thology,underwent DWI at 3.0T before operation.DWI based on SE-EPI sequence,of which b value was 600 s/mm2 ,included BH-DWI and RTIR-DWI.Apparent diffusion coefficient(ADC)value of pancreatic cancer and mass-like pancreatitis were statis-tically analyzed and compared.Receiver operating characteristic(ROC)curve was used to analyze and compare the effect of two DWI sequences in the differential diagnosis of pancreatic cancer and mass-like pancreatitis.Results:On BH-DWI,ADC value did not have significant difference between pancreatic cancer and mass-like pancreatitis(P >0.05).On RTIR-DWI,ADC value had significant difference between pancreatic cancer and mass-like pancreatitis(P <0.01).On RTIR-DWI,the sensitivity,speci-ficity,positive predictive value and negative predictive value were 86.7%,88.9%,96.3% and 66.7%,respectively,when ADC≥1.33×10-3 mm2/s was used as a cutoff for the differential diagnosis of pancreatic cancer from mass-like pancreatitis.On BH-DWI,the sensitivity,specificity,positive predictive value and negative predictive value was 73.3%,88.9%,95.7% and 50.0%,respectively,when ADC≥ 1.45 × 10-3 mm2/s was used as a cutoff for the differential diagnosis of pancreatic cancer from mass-like pancreatitis.Conclusions:Compared to BH-DWI,RTIR-DWI may be more conducive to the diagnosis and differentiation of pancreatic cancer and mass-like pancreatitis.

ObjectiveTo investigate the significance of cerebrospinal fluid(CSF) S-100B protein and vascular endothelial growth factor (VEGF) levels in the pathogenesis of brain injury of viral encephalitis.MethodsForty-two patients with viral encephalitis (viral encephalitis group) and 40 patients with other disease at the corresponding time period(control group) were involved in this study.CSF (routine,biochemistry and cytology) was detected,and the levels of S-100B protein and VEGF in CSF were detected by ABC-ELISA method.ResultsWhite blood cell count was (0-584) × 106/L in viral encephalitis group,and (0-200) × 106/L in control group.The levels of protein and glucose in CSF had no significant difference between two groups (P＞ 0.05),and the level of chloride in CSF in viral encephalitis group was significantly lower than that in control group[ ( 110.10 ± 31.22 ) mmol/L vs.( 123.80 ± 6.32 ) mmol/L ] (P =0.006).In viral encephalitis group,cytological examination showed that mixed type cytological reaction was in 6 cases (14.3％,6/42).The level of S-100B protein in viral encephalitis group [25.04-47.97 (28.37 ± 6.09) ng/L] was significantly higher than that in control group[ 25.04-29.64(26.03 ± 0.90) ng/L ] (t =2.462,P =0.018).The level of VEGF in viral encephalitis group[88.84～143.77(96.24 ± 13.38) ng/L] was significantly higher than that in control group [89.15～96.18 (90.67 ± 1.71 ) ng/L] (t =2.673,P =0.011 ).ConclusionsThe high levels of S-100B protein and VEGF in CSF could support the viral encephalitis diagnosis.Tracking the levels of S-100B protein and VEGF in CSF dynamically have noticeable effect on checking the condition of viral encephalitis patients.