BACKGROUND: Streptococcus mutans (S. mutans) is generally considered to be the principal aetiological agent for dental caries, thrS gene may relate to the virulence of S. mutans involved in the adherence, acidogenicity and acidodurance.OBJECTIVE: To investigate the conservation status of the thrS gene of S. mutans and to construct the homologous recombinant plasmid.METHODS: Southern Blot was used to analyze the distdbuUon of thrS gene in S. mutans. The upstream and downstream sequences of thrS gene were cloned respectively into multiple cloning sites of suicide plasmid pFW5 to construct the recombinant plasmid,RESULTS AND CONCLUSION: The thrS gene was conserved in 6 strains of S. mutans in this test. By PCR analysis and enzyme digesting, it was confirmed that S. mutans thrS gene homologous recombinant plasmid was successfully constructed,which can be used in future research of construction of thrS -negative mutans of S. mutans strain UA159.

Objective:To knock the phospho-sugar mutase gene (psm) out of the genome of S. mutans strain UA159 and to construct the mutant will lay a foundation for the study of the function of the psm. Methods:Two upstream and downstream DNA sequences of the psm were selected and cloned respectively into multiple cloning sites I and II of suicide plasmid pFW5 to construct the recombinant plasmid which was confirmed by enzyme digesting and sequencing. The recombinant plasmid was transformed into S. mutans UA159 by natural transformation and antibiotic was used to screen the positive transformants. According to the principle of homologous recombination, allelic exchange between the recombinant plasmid and S. mutans UA159 was achieved. Results:By PCR analysis and sequencing, it was confirmed that the psm of S. mutans UA159 was substituted for resistant gene of spectinomycin. Conclusion: The psm-negative mutants of S. mutans UA159 is successfully constructed.

Objective To investigate the changing characteristics and rules of the different period of articular cartilage in rats after dorsal rhizotomy, and to verify the partial sensory disturbance can cause articular cartilage injury. Methods Thirty-three ten-month-old SPF male Wistar rats were randomly divided into experimental group ( n=18 ) and control group ( n=15) .The rats in the experimental group were sectioned the L3、L4 dorsal roots in the right.The rats in the control group were only incised the skin and paravertebral muscles.To observe the behavior changes in rats.At 2, 6 and 10 weeks, the specimens of the right hind lower end of femur were drawn out to make paraffin sections, then HE staining and Safranin O/Fast Green staining.The changes and characteristics of the morphology of the articular cartilage was observed.Results With the prolonging of period, the right hind limb of the experimental rats through the change process of transient paralysis, coordination of movement disorders and active movement.In the experimental group, the patellar surface of right hind femur gradually became shallow and wide.The articular cartilage underwent rough, cells disorganizated, cells decreased, and duplicated drifted and interrupted tide line. The ratios of ACC/TAC of the experimental group were gradually high(t=5.25~8.13,P <0.05).Conclusion Dorsal rhizotomy can cause injury and degenerative changes in articular cartilage.

Objective To investigate the effects of tirofiban application time on middle-term clinical prognosis in patients with acute ST segment elevation myocardial infarction (STEMI)treated by primary percutsneous coronary intervention (PCI). Methods The study of tirofiban was carried out in 50 patients with STEM[in cardiology department from January to December 2006. Twenty-nine patients were randomized to receive tirofiban after PCI for 24 - 36 hours(short time group, STG) and 21 patients for 48 - 72 hours (long time group,LTG). Clinical characteristics, angiography data, main adverse cardiovascular events (MACE) and coronary restenosis rate in 6-month follow-up of the two groups were. compared. Results Follow-up data showed that there was less intractable angina pectoris (14.3% vs 24.1%, P< 0.05) in LTG. But there was no significant difference in coronary restenosis rate between two groups. Conclusion Long time application of tirofiban following PCI in patients with STEMI could improve middle-term clinical prognosis by alleviating the incidence of intractable angina pectoris.

Objective:To investigate the reproductive toxicity of cadmium sulfide nanoparticles (Nano-CdS) with different particle sizes on male mice.Methods:In January 2019, 30 SPF grade male mice were randomly divided into a control group, an experimental group[CdS Ⅰ group (particle size approximately 5 nm), and a CdS Ⅱ group (particle size approximately 50 nm) ], with 10 mice in each group. The experimental group was orally gavaged with 100 mg/kg, once a day, while the control group was gavaged with an equal volume of physiological saline for 45 consecutive days. After 45 days, levels of cadmium accumulation in testis were determined directly by AAS, deformity and testicular histopathological changes were also observed. Serum testosterone levels were measured by enzyme-linked immunosorbentassay (ELISA), expression levels of P450scc, 17β-HSD and P450c17 mRNA were determined by real-time PCR. P450c17 protein was determinated by Western Blot.Results:The histopathological results showed that the testes of the experimental group mice showed varying degrees of damage; Ultrastructural observation showed that the ultrastructure of mouse testicular cells in each experimental group showed varying degrees of mitochondrial expansion and disappearance of cristae, as well as irregular nuclear membranes. The degree of damage in CdS Ⅰ group was milder than that in CdS Ⅱ group. Compared with the control group, the cadmium content in the testes of the CdS Ⅰ and CdS Ⅱ groups significantly increased ( P=0.001, 0.001), and the CdS Ⅱ group was higher than the CdS Ⅰ group ( P=0.001). Compared with the control group, the levels of testosterone in the CdS Ⅰ and CdS Ⅱ groups decreased with statistical significance ( P=0.001, 0.001). Real time fluorescence quantitative PCR results showed that compared with the control group, the experimental group's P450scc, 17β-HSD. The expression levels of 17β-HSD and P450c17 mRNA were significantly reduced, with statistically significant differences ( P=0.001, 0.001, 0.001), and CdS Ⅱ group 17β-HSD. The expression levels of 17β-HSD and P450c17 mRNA were significantly lower than those of CdS Ⅰ group ( P=0.001, 0.036). The Western Blot assay results showed that the expression levels of P450c17 protein in the testes of CdS Ⅰ and CdS Ⅱ groups of mice were significantly reduced, with statistical significance ( P=0.001, 0.001) ; And the CdS Ⅱ group was significantly lower than the CdS Ⅰ group ( P=0.001). According to Spearman correlation analysis, testosterone levels are correlated with P450scc, P450c17, 17β-HSD mRNA. There is a highly positive correlation between 17β-HSD mRNA levels, with statistically significant differences ( rs=0.88, 0.80, 0.70, P=0.001, 0.001, 0.004) . Conclusion:Nano cadmium sulfide may induce reproductive toxicity by reducing the expression levels of key enzyme genes and enzyme protein activity in testosterone and its synthesis in mice, and the CdS Ⅱ group has a stronger toxic effect.

Objective:To investigate the reproductive toxicity of cadmium sulfide nanoparticles (Nano-CdS) with different particle sizes on male mice.Methods:In January 2019, 30 SPF grade male mice were randomly divided into a control group, an experimental group[CdS Ⅰ group (particle size approximately 5 nm), and a CdS Ⅱ group (particle size approximately 50 nm) ], with 10 mice in each group. The experimental group was orally gavaged with 100 mg/kg, once a day, while the control group was gavaged with an equal volume of physiological saline for 45 consecutive days. After 45 days, levels of cadmium accumulation in testis were determined directly by AAS, deformity and testicular histopathological changes were also observed. Serum testosterone levels were measured by enzyme-linked immunosorbentassay (ELISA), expression levels of P450scc, 17β-HSD and P450c17 mRNA were determined by real-time PCR. P450c17 protein was determinated by Western Blot.Results:The histopathological results showed that the testes of the experimental group mice showed varying degrees of damage; Ultrastructural observation showed that the ultrastructure of mouse testicular cells in each experimental group showed varying degrees of mitochondrial expansion and disappearance of cristae, as well as irregular nuclear membranes. The degree of damage in CdS Ⅰ group was milder than that in CdS Ⅱ group. Compared with the control group, the cadmium content in the testes of the CdS Ⅰ and CdS Ⅱ groups significantly increased ( P=0.001, 0.001), and the CdS Ⅱ group was higher than the CdS Ⅰ group ( P=0.001). Compared with the control group, the levels of testosterone in the CdS Ⅰ and CdS Ⅱ groups decreased with statistical significance ( P=0.001, 0.001). Real time fluorescence quantitative PCR results showed that compared with the control group, the experimental group's P450scc, 17β-HSD. The expression levels of 17β-HSD and P450c17 mRNA were significantly reduced, with statistically significant differences ( P=0.001, 0.001, 0.001), and CdS Ⅱ group 17β-HSD. The expression levels of 17β-HSD and P450c17 mRNA were significantly lower than those of CdS Ⅰ group ( P=0.001, 0.036). The Western Blot assay results showed that the expression levels of P450c17 protein in the testes of CdS Ⅰ and CdS Ⅱ groups of mice were significantly reduced, with statistical significance ( P=0.001, 0.001) ; And the CdS Ⅱ group was significantly lower than the CdS Ⅰ group ( P=0.001). According to Spearman correlation analysis, testosterone levels are correlated with P450scc, P450c17, 17β-HSD mRNA. There is a highly positive correlation between 17β-HSD mRNA levels, with statistically significant differences ( rs=0.88, 0.80, 0.70, P=0.001, 0.001, 0.004) . Conclusion:Nano cadmium sulfide may induce reproductive toxicity by reducing the expression levels of key enzyme genes and enzyme protein activity in testosterone and its synthesis in mice, and the CdS Ⅱ group has a stronger toxic effect.

Objective:To evaluate the role of sphingosine-1-phospho-1 receptor 1 (S1PR1) in remifentanil-induced hyperalgesia in rats with incisional pain and the relationship with KCNQ2/3 potassium channels in the dorsal root ganglia (DRG).Methods:Forty-eight male Sprague-Dawley rats with successful caudal vein catheterization, aged 2-3 months, weighing 260-280 g, were divided into 6 groups ( n=8 each) using a random number table method: control group (group C), S1PR1 inhibitor group (FTY720) group (group F), remifentanil group (group R), remifentanil + S1PR1 inhibitor (FTY720) group (group RF), remifentanil + incision pain group (group RI) and remifentanil + incision pain + S1PR1 inhibitor (FTY720) group (group RIF). In group C, normal saline 0.1 ml· kg -1·min -1 was intravenously infused for 60 min. In group F, FTY720 3 nmol was intrathecally injected at 10 min before normal saline injection, and 0.1 ml · kg -1·min -1 normal saline was infused into the caudal vein for 60 min. Remifentanil 1.0 μg· kg -1·min -1 was infused for 60 min through the caudal vein in group R. In RF group, FTY720 (3 nmol) was intrathecally injected, and 10 min later remifentanil 1.0 μg· kg -1·min -1 was infused via the caudal vein for 60 min. The incisional pain model was established, and remifentanil 1.0 μg· kg -1·min -1 was infused via the caudal vein for 60 min in RI group. In RIF group, FTY720 3 nmol was intrathecally injected at 10 min before remifentanil infusion, then the incisional pain model was developed, and remifentanil 1.0 μg· kg -1·min -1 was infused via the caudal vein at the same time for 60 min. The mechanical paw withdraw threshold (MWT) and thermal paw withdraw latency (TWL) were measured at 24 h before remifentanil or normal saline infusion (T 0) and 2, 6, 24 and 48 h after remifentanil or normal saline infusion (T 1-4). The rats were sacrificed after the last measurement of pain threshold, and the L 4-6 segments of the DRG were taken for determination of the expression of S1PR1, KCNQ2 and KCNQ3 protein and mRNA in the DRG by Western blot and real-time polymerase chain reaction. Results:Compared with group C, the MWT was significantly decreased, and the TWL was shortened at T 1-4, the expression of S1PR1 protein and mRNA in the DRG was up-regulated, the expression of KCNQ2 and KCNQ3 protein and mRNA in the DRG was down-regulated ( P<0.05), and no significant change was found in each parameter in R and RI groups ( P>0.05). Compared with group R, the MWT was significantly decreased, and the TWL was shortened at T 1-4, the expression of S1PR1 protein and mRNA in the DRG was up-regulated, and the expression of KCNQ2 and KCNQ3 protein and mRNA in the DRG was down-regulated in group RI, and the MWT was significantly increased, and the TWL was prolonged at T 1-4, the expression of S1PR1 protein and mRNA in the DRG was down-regulated, and the expression of KCNQ2 and KCNQ3 protein and mRNA in the DRG was up-regulated in group RF ( P<0.05). Compared with group RI, the MWT was significantly increased, and the TWL was prolonged at T 1-4, the expression of S1PR1 protein and mRNA in the DRG was down-regulated, and the expression of KCNQ2 and KCNQ3 protein and mRNA in the DRG was up-regulated in group RIF ( P<0.05). Conclusions:S1PR1 is involved in the process of remifentanil-induced hyperalgesia in rats with incisional pain, which is related to the inhibition of KCNQ2/3 potassium channel expression in the DRG.

Objective: To survey the conduction and evaluate the effectiveness of extracorporeal membrane oxygenation (ECMO) therapy in pediatric intensive care unit (PICU) in China mainland. Methods: In a questionnaire-based survey, we retrospectively reviewed the application of ECMO in children's hospital and general hospital in China mainland to summarize and analyze the categories of diseases and prognosis of children treated with ECMO therapy. Results: By December 31, 2017, a total of 23 hospitals using ECMO, including 22 tertiary referral hospitals and 1 secondary hospital, among which 16 were children′s hospitals and 7 were general hospitals. Thirty-seven ECMO equipment was available. A total of 518 patients treated with ECMO, within whom 323 (62.4%) successfully weaned from ECMO and 262 (50.6%) survived to discharge. Among 375 pediatric patients, 233 (62.1%) were successfully weaned from ECMO and 186 (49.6%) survived to discharge. Among 143 newborn patients, 90 (62.9%) successfully weaned from ECMO, 76 (53.1%) survived to discharge. ECMO was applied in veno-arterial (VA) mode to 501 (96.7%) patients, veno-venous (VV) mode to 14 (2.7%) patients, and VV-VA conversion mode to 3 (0.6%) patients. Sixty-nine patients required extracorporeal cardiopulmonary resuscitation (ECPR), including 20 newborn patients (29.0%) and 38 pediatric patients (71.0%), who were all with cardiovascular disease. Neonatal respiratory distress syndrome (26/61), persistent pulmonary hypertension of the newborn (PPHN) (12/61), and meconium aspiration syndrome (MAS) (11/61) are the most common pulmonary diseases in newborn patients; among whom, infants with PPHN had highest survival rate (10/12), followed by MAS (9/11). Among newborn patients with cardiovascular diseases, those who admitted were after surgery for congenital cardiac disease were the most common (54/82), while those with septic shock had the highest survival rate (2/3). In pediatric pulmonary diseases, acute respiratory distress syndrome was the most common (42/93), while plastic bronchitis was with the highest survival rate (4/4), followed by viral pneumonia (13/16). Among pediatric cardiovascular diseases, congenital cardiac defect was the most common (124/282), while fulminant myocarditis had the highest survival rate (54/77). Conclusion The application of ECMO as a rescue therapy for children with severe cardiopulmonary failure has dramatically developed in China mainland.

Objective: To explore the effect of distance education in the teaching mode of medical clinical skills, and to provide theoretical and practical basis for finding a better teaching mode to promote the combination of theory and practice. Methods: A total of 172 trainees of clinical skill training in the training center were divided into the control group and the observation group (the observation group was divided into pre-class long-distance group, in-class long-distance group, after-class long-distance group). There were 43 trainees in each group. The control group adopted traditional teaching methods in clinical basic skills courses, and the observation group adopted mixed teaching mode under long-distance education. The satisfaction of teachers and students under the two teaching modes were investigated, and the performance of each group of students in skills, human-computer dialogue, and team first aid comprehensive test were compared. Results: There were significant differences in teacher satisfaction between the control group and the pre-class and in-class distance groups (χ² = 15.315, P = 0.000). There were significant differences in student satisfaction between the control group and the pre-class and in-class distance groups in terms of interactive participation, liveliness, interesting training skills and teaching level (χ²=4.497-17.153, P = 0.000-0.034). The results of each group reached the expected teaching goals. There were significant differences between the skill test score control group and the pre-class and in-class remote groups (t=25.357, 14.712, all P =0.000). There were significant differences between the human-machine dialogue test control group and the three observation groups (t=14.561, 19.420, 3.821, all P =0.000). There were significant differences between the team emergency comprehensive test control group and the three observation groups (t=14.561, 19.420, 3.821, all P=0.000). Conclusions Distance education has flexible in the mixed teaching mode of clinical skills, especially in pre-class and in-class. It can rapidly improve the basic clinical skills, theoretical knowledge and team mobilization ability of students. It is one of the best bridges to communicate their clinical theory and practice.

Nonalcoholic fatty liver disease (NAFLD) is characterized by hepatic steatosis and insulin resistance and there are currently no approved drugs for its treatment. Hyperactivation of mTOR complex 1 (mTORC1) and subsequent impairment of the transcription factor EB (TFEB)-mediated autophagy-lysosomal pathway (ALP) are implicated in the development of NAFLD. Accordingly, agents that augment hepatic TFEB transcriptional activity may have therapeutic potential against NAFLD. The objective of this study was to investigate the effects of nuciferine, a major active component from lotus leaf, on NAFLD and its underlying mechanism of action. Here we show that nuciferine activated ALP and alleviated steatosis, insulin resistance in the livers of NAFLD mice and palmitic acid-challenged hepatocytes in a TFEB-dependent manner. Mechanistic investigation revealed that nuciferine interacts with the Ragulator subunit hepatitis B X-interacting protein and impairs the interaction of the Ragulator complex with Rag GTPases, thereby suppressing lysosomal localization and activity of mTORC1, which activates TFEB-mediated ALP and further ameliorates hepatic steatosis and insulin resistance. Our present results indicate that nuciferine may be a potential agent for treating NAFLD and that regulation of the mTORC1-TFEB-ALP axis could represent a novel pharmacological strategy to combat NAFLD.