Objective:To investigate the functional mechanism of circular RNA signal-induced proliferation-associated gene 1(circSIPA1L1) on proliferation, migration and invasion of renal cell carcinoma cells, as well as to explore its mechanism.Methods:The study was completed between January 2019 and December 2019. Bioinformatics was used to analyze the expression of circular RNA(circRNA), circSIPA1L1 in renal cancer tissue and the information of circSIPA1L1. The expression of circSIPA1L1 mRNA, miR-22-3p in renal cancer tissues and renal cancer cells was detected by RT-qPCR. The circSIPA1L1 interference vector negative control (si-NC group), circSIPA1L1 interference vector (si-circSIPA1L1 group), si-circSIPA1L1+ miR-22-3p suppression vector plasmid negative control (anti-miR-NC group), si-circSIPA1L1 + miR-22-3p inhibition vector plasmid (anti-miR-22-3p group) were transfected into A498 and OSRC2 cells respectively. Dual luciferase reporter gene experiment was used to verify the targeting relationship. Clone formation experiment and Transwell chamber were used to detect cell proliferation, migration and invasion. The xenograft model was established by subcutaneous injection of A498/sh-circSIPA1L1 or A498/sh-NC (2×10 6 in 0.2 ml PBS/mice) on the right back of nude mice, and nude mice were divided into sh-circSIPA1L1 group and sh-NC group. Nude mice tumor formation experiments were used to detect tumor formation ability. Results:The expression of circSIPA1L1 mRNA in adjacent tissues and renal cancer tissues were (1.09±0.44) and (3.89±1.35) respectively. The expression of miR-22-3p were (1.02±0.30) and (0.44±0.19)respectively. The difference of the expression of circSIPA1L1 mRNA and miR-22-3p in kidney cancer tissue and adjacent tissues were statistically significant ( P<0.05). Compared with normal kidney cell KiMA, the expression of circSIPA1L1 mRNA in renal cancer cells A498 and OSRC2 was increased, and the expression of miR-22-3p was decreased ( P<0.05). The cell clone number of A498 and OSRC2 in the si-circSIPA1L1 group (130.67±15.04, 99.00±14.80) was lower than that in the si-NC group (314.33±29.57, 234.67±21.50), the number of cell migration (108.33±17.01, 85.67±11.93) was lower than si-NC group (265.00±20.00, 210.33±18.58), cell invasion number (84.00±12.00, 66.00±10.15) was lower than si-NC group (210.33±18.58, 173.00±17.52), and the differences were all statistically significant ( P< 0.05). CircSIPA1L1 targets and negatively regulates miR-22-3p expression. The cell clone number of A498 and OSRC2 in the si-circSIPA1L1+ anti-miR-22-3p group (234.20±21.90, 185.06±20.72) was higher than that in the si-circSIPA1L1+ anti-miR-NC group (134.65±26.55, 106.14±16.38), the migration cell number (187.02±23.54, 117.86 ±15.09) was higher than that of the si-circSIPA1L1+ anti-miR-NC group (110.59±12.12, 91.70±14.83), and the number of cell invasion (168.23±11.69, 103.70±9.23) was higher than that of the si-circSIPA1L1+ anti-miR-NC group (90.46±11.53, 61.35±9.10). The differences were all statistically significant ( P<0.05). The tumor volumes of nude mice in the sh-NC group and sh-circSIPA1L1 group on day 35 were (578.65±68.67) mm 3 and (242.56±42.35) mm 3 respectively, the tumor weights of nude mice were (0.68±0.06) g and (0.38±0.04) g respectively, the differences were statistically significant ( P<0.05). Conclusions:CircSIPA1L1 can promote the deterioration of renal cancer, promote the proliferation, migration, invasion of cancer cells and tumor growth. The mechanism of action is related to the direct targeting of miR-22-3p.

Objective To construct a naive human Fab fragment phage display library,from which the anti-gp96 antibodies may be panned by the gp96 purified from the tissue of urothelial carcinoma in the urinary bladder and provide a basis to new therapy for the malignant tumors.Methods Peripheral blood lymphocytes were isolated from 800 ml of blood,which was obtained from four healthy blood donors.The heavy chain Fd and light chain cDNA synthesized from the total RNA of lympbocytes were amplified by PCR with variable regions 5' and 3' primers of heavy and light chain, and the amplification products were ligated into the phagemid vector pComb3, then the ligated sample was transformed into competent E.coli XL1-Blue by electroperation.The transformed cells were infected with VCSM13 helper phage to yield recombinant phage antibody Fabs.The phagemids abstracted from amplified E.coli were cut with endonucleases such as Sac Ⅰ,Xba Ⅰ,Xho Ⅰ and Spe Ⅰ, and both the phage antibody Fabs and phage-raids abstracted from amplified E.coil were amplified by PCR to monitor the insertion of the genes of light chain or heavy chain Fd fragment.The gp96 purified from the urothelial carcinoma tissue of the bladder by affinity chromatog-raphy on eoncanavalin-A sephnrose and DEAE-sephnrose ion exchange chromatography were utilized as antigens to process three rounds of panning to the original Fab antibody library.Results The quantity of total RNA and cDNA were qualified.By combination of light chain and heavy chain genes, an antibody library containing 6.6×106 clones was obtained, and both the cutting of enzymes and PCR showed that there were the genes of light chain or heavy chain Fd fragment in the phagemids.The gp96 protein was obtained from urothelial carcinoma tissue in the urinary bladder.After having been panned by gp96, the original antibody library gained enrichment by 68 times.Conclusion Utilizing the technology of phage surface display, specific antibody can be gained from the human naive Fab phage display library,which can be used for immunological therapy for tumors.

Objective To discuss the changes of correlative ultrasonic parameter index of normal hips and abnormal hips with developmental dislocation of the hip (DDH) in infants of different months in order to provide objective information for the diagnosis. Methods Three-hundred and seventy-eight normal hips and 244 hips with DDH among 622 hips of 311 infants were detected by ultrasonography(US). The morphology and structure information of hips were observed, and the values of ultrasonic parameter index,including angle α,angle 3, acetabular index( AI), femoral head percentage of cover(FHC) of normal hips and abnormal hips were measured. The values of each parameter index were collected by being divided into different groups (3 months a group) ,then the correlation was analyzed. Results Morphology and structure,position relation between femoral head and acetabulum of the hips were demonstrated by US. Normal or abnormal hips,the degrees of abnormal hips and the types of hips could be judged according to the findingsof US. Analysis of values of parameter index of normal hips:①There was significantly statistical significance in the values of ultrasonic parameter index, such as angle α, angle β, AI, FHC of normal hip between the groups of different age (P＜0.01). ②There was correlation between the age and the values of each parameter index, among which angle α, FHC had positive correlations with age ( r = 0. 537, 0. 554,respectively ) while angle β and Al negative correlations ( r = -0. 465, -0.424, respectively ). ③There was correlation between the values of different parameter index. Both angle β and AI had negative correlation with angle α,among which the latter correlation was closely ( r = - 0. 794). No statistical significance was found between the ultrasonic values of each group under different ages of different type abnormal hips( P ＞0.05) ,but closely negative correlations still existed between angle α and AI. ConclusionsUS can be viewed as an early definite and a screening method of diagnosing DDH.For older infants (above 6 months) it will be more accurate to analyze the ultrasonic parameter index together with the age of infants.

Objective To investigate whether activation of mitochondrial acetal dehydrogenase 2(ALDH2)alleviates hypoxic pulmonary hypertension by regulating the SIRT1/PGC-1α signaling pathway.Methods Thirty 8-week-old C57 BL/6 mice were randomized into control,hypoxia,and hypoxia+Alda-1(an ALDH2 activator)group(n=10),and the mice in the latter two groups,along with 10 ALDH2 knockout(ALDH2-/-)mice,were exposed to hypoxia(10%O2,90%N2)with or without daily intraperitoneal injection of Alda-1 for 4 weeks.The changes in right ventricular function and pressure(RVSP)of the mice were evaluated by echocardiography and right ventricular catheter test,and pulmonary artery pressure was estimated based on RVSP.Pulmonary vascular remodeling,right ventricular injury,myocardial α-SMA expression,distal pulmonary arteriole muscle normalization,right ventricular cross-sectional area,myocardial cell hypertrophy,and right cardiac hypertrophy index were assessed with HE staining,immunofluorescence staining and WGA staining,and the expressions of ALDH2,SIRT1,PGC-1α,P16INK4A and P21CIP1 were detected.In pulmonary artery smooth muscle cells with hypoxic exposure,the effect of Alda-1 and EX527 on cell senescence and protein expressions was evaluated using β-galactose staining and Western blotting.Results The wild-type mice with hypoxic exposure showed significantly increased RVSP,right ventricular free wall thickness and myocardial expressions of P16INK4A and P21CIP1,which were effectively lowered by treatment with Alda-1 but further increased in ALDH2-/-mice.In cultured pulmonary artery smooth muscle cells,hypoxic exposure significantly increased senescent cell percentage and cellular expressions of P16INK4A and P21CIP1,which were all lowered by treatment with Alda-1,but its effect was obviously attenuated by EX527 treatment.Conclusion ALDH2 alleviates hypoxia-induced senescence of pulmonary artery smooth muscle cells by upregulating the SIRT1/PGC-1α signaling pathway to alleviate pulmonary hypertension in mice.

Objective To investigate whether activation of mitochondrial acetal dehydrogenase 2(ALDH2)alleviates hypoxic pulmonary hypertension by regulating the SIRT1/PGC-1α signaling pathway.Methods Thirty 8-week-old C57 BL/6 mice were randomized into control,hypoxia,and hypoxia+Alda-1(an ALDH2 activator)group(n=10),and the mice in the latter two groups,along with 10 ALDH2 knockout(ALDH2-/-)mice,were exposed to hypoxia(10%O2,90%N2)with or without daily intraperitoneal injection of Alda-1 for 4 weeks.The changes in right ventricular function and pressure(RVSP)of the mice were evaluated by echocardiography and right ventricular catheter test,and pulmonary artery pressure was estimated based on RVSP.Pulmonary vascular remodeling,right ventricular injury,myocardial α-SMA expression,distal pulmonary arteriole muscle normalization,right ventricular cross-sectional area,myocardial cell hypertrophy,and right cardiac hypertrophy index were assessed with HE staining,immunofluorescence staining and WGA staining,and the expressions of ALDH2,SIRT1,PGC-1α,P16INK4A and P21CIP1 were detected.In pulmonary artery smooth muscle cells with hypoxic exposure,the effect of Alda-1 and EX527 on cell senescence and protein expressions was evaluated using β-galactose staining and Western blotting.Results The wild-type mice with hypoxic exposure showed significantly increased RVSP,right ventricular free wall thickness and myocardial expressions of P16INK4A and P21CIP1,which were effectively lowered by treatment with Alda-1 but further increased in ALDH2-/-mice.In cultured pulmonary artery smooth muscle cells,hypoxic exposure significantly increased senescent cell percentage and cellular expressions of P16INK4A and P21CIP1,which were all lowered by treatment with Alda-1,but its effect was obviously attenuated by EX527 treatment.Conclusion ALDH2 alleviates hypoxia-induced senescence of pulmonary artery smooth muscle cells by upregulating the SIRT1/PGC-1α signaling pathway to alleviate pulmonary hypertension in mice.

Objective:To explore the standardized performance of a FISH probe before clinical detection.Methods:The probe sensitivity and specificity of ETV6/RUNX1 were analyzed via interphase and metaphase FISH in 20 discarded healthy bone marrow samples. The threshold system of the probe was established using an inverse beta distribution, and an interpretation standard was established. Finally, a parallel-controlled polymerase chain reaction detection study was conducted on 286 bone marrow samples from patients at our hospital. The clinical sensitivity, specificity, and diagnostic coincidence rate of ETV6/RUNX1 FISH detection were analyzed, and the diagnostic consistency of the two methods was analyzed by the kappa test.Results:The probe sensitivity and specificity of the ETV6/RUNX1 probe were 98.47% and 100%, respectively. When 50, 100, and 200 cells were counted, the typical positive signal pattern cutoffs were 5.81%, 2.95%, and 1.49%, respectively, and the atypical positive signal pattern cutoffs were 13.98%, 9.75%, and 6.26%, respectively. The clinical sensitivity of FISH was 96.1%, clinical specificity was 99.6%, diagnostic coincidence rate was 99.00%, diagnostic consistency test kappa value was 0.964, and P value was <0.001.Conclusion:For FISH probes without a national medical device registration certificate, standardized performance verification and methodology performance verification can be performed using laboratory developed test verification standards to ensure a reliable and accurate reference basis for clinical diagnosis and treatment.

【Objective】 To investigate the feasibility of using surface electromyography （SEMG） for the detection of abnormal muscle response （AMR） in patients with hemifacial spasm （HFS）. 【Methods】 We retrospectively reviewed the clinical data of HFS patients who underwent microvascular decompression （MVD） in our hospital between June 2019 and December 2020. Patients who received both surface electrode （preoperative） and needle electrode （intraoperative） detection of AMR were included. SEMG recorded from two stimulation-recording sites， namely， zygomatic-mentalis and mandibular marginal-orbicularis oculi， was selected for analyzing the characteristics of AMR. The positive rates of AMR detected by these two kinds of electrodes were comprehensively compared. 【Results】 Totally 77 patients were included in this study. When detected with surface electrodes， the positive rate， latency and amplitude of AMR recorded at zygomatic-mentalis oculi were 90.9% （70/77）， （10.87±1.86） ms and （202.8±47.4） μV， and at mandibular marginal-orbicularis oculi were 92.2% （71/77）， （10.41±1.83） ms and （211.1±54.1） μV， respectively. AMR was detected in 74 patients （96.1%） with surface electrodes. There was no significant difference in positive rate， latency and amplitude of AMR between these two stimulation-recording methods. When detected with needle electrodes， the positive rate of AMR recorded at zygomatic-mentalis oculi was 98.7% （76/77）， which was significantly higher than the rate 89.6% （69/77） recorded at mandibular marginal-orbicularis oculi （P=0.016）. The latency and amplitude of AMR recorded at zygomatic-mentalis were （10.63±1.39） ms and （83.5±27.2） μV， and at mandibular marginal-orbicularis oculi were （10.31±1.18） ms and （58.6±21.4） μV. There was no significant difference in latency between the two stimulation-recording methods， but the amplitude recorded at mandibular marginal-orbicularis oculi was significantly lower （P=0.041）. AMR was detected in 76 patients （98.7%） with needle electrodes. There was no significant difference in the detection rate of AMR between surface electrodes and needle electrodes （P=0.500）， the results were moderately consistent （Kappa=0.490， P<0.001）. 【Conclusion】 The detection efficiency of surface electrodes for AMR is similar to that of needle electrode. With its non-invasive characteristic， the surface electrode can be routinely used for electrophysiological evaluation of HFS.