AIM:To investigate the mechanism of Piezo1 channel activation promoting the increase in intra-cellular Ca2+ concentration([Ca2+]i)of rat coronary artery smooth muscle cells(CASMCs).METHODS:The primary CASMCs of SD rats were cultured,and the expression and subcellular localization of Piezo1 in the cells were observed by immunofluorescence staining.The Piezo1 and stromal interaction molecule 1(STIM1)in CASMCs were knocked down by siRNA transfection,and the expression levels of the proteins were detected by Western blot.Utilizing laser confocal mi-croscopy,the change of[Ca2+]i in CASMCs was detected by Fluo-4 AM fluorescent probes.RESULTS:It was confirmed by immunofluorescence staining that the expression of Piezo1 existed in primary rat CASMCs.Immunofluorescence staining also showed that Piezo1 was co-located with sarco-/endoplasmic reticulum Ca2+-ATPase 2(SERCA2),mitochondrial outer membrane protein TOM20 and nuclear membrane protein lamin B1.Western blot results showed that the protein expres-sion levels of STIM1 and Piezo1 were significantly down-regulated by siRNA transfection(P<0.05).Compared with con-trol group,Yoda1,the agonist of Piezo1,could increase the extracellular Ca2+ influx of CASMCs(P<0.01).However,the Ca2+ influx mediated by Yoda1 was not affected by the inhibition of L-type calcium channels.Treatment with Yoda1 in-creased the intracellular Ca2+ release of CASMCs(P<0.01).However,inhibition of calcium channels on endoplasmic re-ticulum,ryanodine receptor and inositol 1,4,5-triphosphate receptor,did not affect intracellular Ca2+ release mediated by Yoda1.After the Ca2+ in endoplasmic reticulum was emptied using thapsigargin(TG),Yoda1 also mediated the Ca2+ re-lease of other organelles in CASMCs(P<0.01).After inhibition of L-type calcium channels,treatment with store-operated calcium channel(SOCC)inhibitor BTP2 or knockdown of STIM1 led to the decrease in extracellular Ca2+ influx of CASMCs mediated by Yoda1(P<0.01).Treatment with TG increased the release of Ca2+ from the endoplasmic reticulum of CASMCs after knockdown of Piezo1(P<0.05),but the extracellular Ca2+ influx mediated by TG was not affected.After inhibition of L-type calcium channels and SOCC,knockdown of Piezo1 led to the decreases in intracellular Ca2+ release and extracellular Ca2+ influx induced by Yoda1(P<0.01).CONCLUSION:The Piezo1 agonist orchestrates the influx of extracellular Ca2+ by activating Piezo1 channels on the cell membrane and inducing the indirect activation of SOCC.More-over,it facilitates the release of Ca2+ from organelles.Consequently,these pathways synergistically elevate the[Ca2+]i of rat CASMCs.

AIM:To investigate the influence of fat mass and obesity-associated(Fto)gene on the aberrant contraction of aortic smooth muscle in diabetes mellitus(DM)mice,and to explore the mechanism of Fto gene underlying the calcium regulation.METHODS:Smooth muscle-specific Fto gene knockout(FtoSMKO)mice were generated using Cre-loxP technology.The experiment involved 3 groups of mice:wild-type(WT)group,DM model group and FtoSMKO-DM group,with 15 mice in each group.In DM group and FtoSMKO-DM group,type 1 DM was induced by intraperitoneal injec-tion of streptozotocin.The mice in WT group were injected with equal volume of citric acid-sodium citrate buffer solution.The influences of different drugs on the contraction responses of aortic smooth muscle in mice were analyzed using a multi-myograph system.The expression level of FTO protein in the aortic tissues was detected by Western blot.RESULTS:(1)Compared with WT mice,the expression levels of FTO protein in the aortic tissues of DM mice were significantly in-creased(P＜0.01).(2)The expression level of FTO protein in smooth muscle was significantly decreased after knockout of Fto gene(P＜0.01).Compared with WT group,the mice in DM group exhibited a significant decrease in body weight and a marked increase in fasting blood glucose level(P＜0.05).There were no noticeable differences in body weight or fasting blood glucose level between FtoSMKO-DM group and DM group(P＞0.05).(3)The contraction responses of aortic smooth muscle in DM group were substantially increased by phenylephrine compared with WT group.Specifically,vaso-constriction responses mediated by non-L-type calcium channels and store-operated calcium channels(SOCC)were signifi-cantly enhanced in DM group.In addition,the responses mediated by inositol 1,4,5-trisphosphate receptors(IP3R),which facilitate calcium release from the sarcoplasmic reticulum,were significantly enhanced.However,the responses mediated by caffeine-activated ryanodine receptors(RyR),which also facilitate calcium release from the sarcoplasmic re-ticulum,were significantly inhibited(P＜0.05).(4)Compared with DM group,the phenylephrine-induced contraction re-sponses of aortic smooth muscle in FtoSMKO-DM group were greatly weakened(P＜0.05).In particular,the vasoconstriction responses mediated by non-L-type calcium channels and SOCC in FtoSMKO-DM group were greatly suppressed(P＜0.05),while those mediated by caffeine-activated RyR were dramatically boosted(P＜0.05).However,IP3R-mediated responses were not affected(P＞0.05).CONCLUSION:Smooth muscle-specific Fto gene knockout suppresses contractile hyperre-sponsiveness in the aortic smooth muscle of DM mice,which may be attributed to involvement of FTO protein in calcium regulation in the vascular smooth muscle.

Objective To investigate the progress of two-dimensional echocardiographic parameters in patients with different severity of aortic stenosis.Methods A retrospective analysis was per-formed on 96 patients diagnosed with aortic stenosis with at least 2 times of transthoracic echo-cardiography(interval ≥1 year)in Department of Cardiology,Fourth Medical Center of Chinese PLA General Hospital from March 2017 to December 2023.According to aortic stenosis severity,they were divided into a mild group(72 cases),a moderate group(14 cases)and a severe group(10 cases).Peak pressure gradient(PPG)across aortic valve,Vmax,mean aortic valve pressure gradient(ΔPm),pulmonary artery systolic pressure(PASP)were collected,and the changes and annual progress of these echocardiographic parameters at baseline and before and after follow-up were analyzed.Results The values of IVST,LVPWT,Vmax,aortic valve PPG and ΔPm were sig-nificantly increased in the mild,moderate and severe stenosis groups in turn(P＜0.05,P＜0.01).The values of Vmax,PPG and ΔPm were significantly lower in the mild stenosis group than the moderate and severe stenosis groups,and the LVPWT value was obviously lower in the mild ste-nosis group than the severe stenosis group(P＜0.05).The aortic valve PPG and ΔPm values at follow-up were significantly higher than those before the follow-up in the three stenosis groups(P＜0.05,P＜0.01).After follow-up,the Vmax values in mild and moderate stenosis groups were notably higher than before(P＜0.01).The PASP value at follow-up was significantly higher than before in the severe stenosis group(P＜0.05).The annual progression rate of Vmax,PASP,LVEF were gradually increased in the mild,moderate,and severe stenosis groups(P＞0.05).The annual progression rate of ΔPm was gradually increased in the three groups in turn(2.30±1.77 mm Hg/year vs 2.40±1.18 mm Hg/year vs 6.08±1.70 mm Hg/year,P＜0.05).Conclusion As the severity of baseline aortic stenosis increases,obvious changes are observed in cardiac structure and function.Before and after follow-up,the serious the aortic stenosis severity is,the faster the annual progression rates of Vmax,PPG,LVEF and PASP are.

OBJECTIVE: To carry out prenatal diagnosis for a fetus with normal ultrasonographic finding at 20 weeks' gestation but a copy number variant(CNV) of 13q indicated by non-invasive prenatal test (NIPT). METHODS: Karyotyping analysis and chromosomal CNV assay were carried out on the amniotic fluid sample. Parental peripheral blood sample was collected for chromosomal analysis. Detailed fetal ultrasound scan was carried out to rule out structural abnormalities of the fetus. RESULTS: The fetus was detected with a heterozygous 10.14 Mb deletion at 13q21.1q21.32, which has originated from the phenotypically normal mother. No apparent karyotypic abnormality was detected in the fetus and its parents. No ultrasonic abnormality was found in the fetus. CONCLUSION Both the fetus and its mother have carried a heterozygous 10.14 Mb deletion at 13q21.1q21.32 and presented normal phenotypes.Combined with literature review, the segmental deletion was judged to be a benign variant.
Female ; Genetic Counseling ; Humans ; Karyotyping ; Pedigree ; Pregnancy ; Prenatal Diagnosis ; Ultrasonography, Prenatal

Objective:To compare the effectiveness and recurrence rate of different types of mesh or without mesh in laparoscopic hiatal hernia repair.Methods:From Jan 2016 to Mar 2022 at the three hospital 90 patients with hiatal hernia, including 26 cases without mesh, 29 cases using synthetic mesh, and 35 cases using biological mesh underwent laparoscopic hiatal hernia repair.Results:The surgical procedures was successful in all the 90 cases without conversion to open surgeny. There were no statistically significant differences in operative time, intraoperative blood loss and postoperative hospital stay among the three groups ( P>0.05), and there were statistically significant differences in hospital cost between the group without mesh and synthetic mesh and biological mesh ( P<0.05). Long-term follow-up was achieved in 87 patients, with a follow-up rate of 96.7% (87/90), and a median follow-up time of 44 months. There were no significant differences in the incidence of postoperative complications (diarrhea, dysphagia, abdominal distension, chest pain), recurrence rate of symptoms (acid reflux, heartburn) and patient satisfaction among the three groups ( P>0.05). Conclusion:In laparoscopic hiatal hernia repair, the mesh should be carefully selected according to the specific intraoperative situation for a satisfactory clinical efficacy.

Objective:To explore the risk factors of chronic postoperative inguinal pain for laparoscopic trans-abdominal preperitoneal hernia repair and establish a nomogram prediction model for it.Methods:The clinical data of 576 patients who underwent laparoscopic trans-abdominal preperitoneal hernia repair for inguinal pain at the First Hospital of Lanzhou University from January 2015 to December 2020 were analyzed retrospectively. According to different postoperative outcomes, patients were divided into chronic pain group ( n=54) and non-chronic pain group ( n=522), compared two groups of patients in the material, including gender, age, BMI, smoking history, history of drinking, hypertension, diabetes, chronic bronchitis, abdominal surgery history, history of inguinal hernia, hernia type, the hernial sac size, prophylactic use of antibiotics, VAS score, mesh fixation techniques, operation time, length of stay. Measurement data with normal distribution were expressed as ( ± s) and independent sample t test was used for comparison between groups. Measurement data with skewed distribution were expressed as M( Q1, Q3), and the Mann-Whitney U test was used for comparision between groups. Chi-square test was used to compare the measurement data of counting data.Multivariate logistic regression was used to analyze the independent risk factors for chronic postoperative inguinal pain. R software was used to establish the drawing of the nomogram prediction model, and the consistency index, calibration chart and area under the receiver operating characteristic curve was used to evaluate the predictive ability of the nomogram prediction model. Results:According to the results of the Logistic regression analysis, age≤45 years ( OR=2.202, 95% CI: 1.080-4.491), BMI≥24 kg/m 2 ( OR=2.231, 95% CI: 1.204-4.134), hernial sac≤5 cm ( OR=2.623, 95% CI: 1.309-5.257), recurrent hernia ( OR=2.769, 95% CI: 1.118-6.860), preoperative pain ( OR=4.121, 95% CI: 2.004-8.476), suture fixation ( OR=2.204, 95% CI: 1.151-4.219)and Postoperative acute pain (VAS>3) ( OR=5.814, 95% CI: 2.532-13.350) were independent risk factors for chronic postoperative inguinal pain ( P<0.05). Based upon the above independent risk factors, the nomogram prediction model was established and verified. The area under the curve of the nomogram prediction model was 0.779 (95% CI: 0.718-0.840, P<0.01). After internal verification, the concordance index value of the prediction model was 0.779. Conclusion:age≤45 years, BMI ≥24 kg/m 2, hernial sac≤5 cm, recurrent hernia, preoperative pain, suture fixation and Postoperative acute pain (VAS>3) are independent risk factors for chronic postoperative inguinal pain for laparoscopic trans-abdominal preperitoneal hernia repair, the nomogram prediction model has a good accuracy and discrimination with a high value of clinical application.

Objective:To investigate the application value of laparoscopy for stage Ⅳ gastric cancer after transformation therapy.Methods:The clinical data of 7 patients with advanced gastric cancer who underwent laparoscopic surgery after transformation therapy in the First Affiliated Hospital of Hebei North University from January 2016 to December 2020 were retrospectively analyzed. Seven patients were diagnosed as stage Ⅳ gastric cancer by computed tomography (CT), positron emission tomography (PET-CT) and laparoscopy, and underwent paclitaxel based conversion therapy. After evaluating the curative effect, they underwent surgical resection. The intraoperative and postoperative conditions and survival rate of the patients were analyzed.Results:After transformation therapy, 6 patients (85.7%) had partial remission, 1 patient (14.3%) had stable disease, and the overall effective rate was 6/7. Laparoscopic-assisted surgery was performed in 3 patients, and 4 patients were converted to laparotomy. The operation time was (297.9±35.6)min, the intraoperative blood loss was (257.0±106.0)ml, the number of dissected lymph nodes was (38.4±9.1), the gastric tube indwelling time was (72.4±9.6)h, the jejunal feeding tube indwelling time was (15.4±5.6)d, and the liquid diet time was (8.6±3.4)d, the length of hospital stay was (17.1±5.5)d. The postoperative complication rate was 3/7, and there were no unplanned secondary operations and deaths. R0 resection was performed in 6 cases, and R1 resection in 1 case. Tumor regression grade (TRG) classification: 5 cases were grade 2 and 2 cases were grade 3. The median progression free survival of the 7 patients was 15.3 months, the median overall survival was 21.6 months, and the 1-year survival rate was 6/7.Conclusions:Laparoscopy has irreplaceable value in the staging and efficacy judgment of transformation therapy for gastric cancer. However, after transformation therapy, the tissue edema increases, the tumor boundary becomes more unclear, and the surgical operation becomes more difficult. Therefore, it is not necessary to force the laparoscopic operation, and switch to laparotomy according to the situation.

OBJECTIVE: To detect variants of IVD gene among 4 neonates with suspected isovalerate acidemia in order to provide a guidance for clinical treatment. METHODS: 111 986 newborns and 7461 hospitalized children with suspected metabolic disorders were screened for acyl carnitine by tandem mass spectrometry. Those showing a significant increase in serum isovaleryl carnitine (C5) were analyzed for urinary organic acid and variants of the IVD gene. RESULTS: Four cases of isovalerate acidemia were detected, which included 2 asymptomatic newborns (0.018‰, 2/111 986) and 2 children suspected for metabolic genetic diseases (0.268‰, 2/7461). The formers had no obvious clinical symptoms. Analysis of acyl carnitine has suggested a significant increase in C5, and urinary organic acid analysis has shown an increase in isovaleryl glycine and 3-hydroxyisovalerate. Laboratory tests of the two hospitalized children revealed high blood ammonia, hyperglycemia, decreased red blood cells, white blood cells, platelets and metabolic acidosis. The main clinical manifestations have included sweaty foot-like odor, feeding difficulty, confusion, drowsiness, and coma. Eight variants (5 types) were detected, which included c.158G>A (p.Arg53His), c.214G>A (p.Asp72Asn), c.548C>T (p.Ala183Val), c.757A>G (p.Thr253Ala) and 1208A>G (p.Tyr403Cys). Among these, c.548C>T and c.757A>G were unreported previously. None of the variants was detected by next generation sequencing of 2095 healthy newborns, and all variants were predicted to be likely pathogenic based on the guidelines from the American College of Medical Genetics and Genomics. CONCLUSION The incidence of isovalerate acidemia in Liuzhou area is quite high. Screening of metabolic genetic diseases is therefore recommended for newborns with abnormal metabolism. The discovery of novel variants has enriched the mutational spectrum of the IVD gene.
Infant, Newborn ; Child ; Humans ; Acidosis ; Carnitine ; Erythrocytes ; High-Throughput Nucleotide Sequencing

Objective:To investigate the effect of cyclin D1 (CCND1) negatively regulated by miRNA-541 (miR-541-5p) on the proliferation and migration of colon cancer cells as well as its related mechanism.Methods:Expression levels of miR-541-5p in colon cancer cell lines HT29, SW480, SW620, HCT116 and enterocyte line HIEC of the normal people as well as cancer tissues and pericarcinomatous normal tissues of 112 patients undergoing the colon cancer surgery from the First Affiliated Hospital of Hebei North University between April 2017 and March 2020 were detected by using quantitative real-time polymerase chain reaction(qRT-PCR). The potential target gene of miR-541-5p was predicted by using TargetScan, and was verified by using dual luciferase reporter gene assay, qRT-PCR and Western blot. Expression level of CCND1 was detected in colon cancer cell lines and tissues. Cells with the lowest expression level of miR-541-5p were divided into miR-NC group (the transfected control plasmid), miR-541-5p group (the transfected miR-541-5p mimics), miR-541-5p+CCND1 group (the co-transfected miR-541-5p mimics and CCND1). Effect of miR-541-5p and CCND1 on proliferation and migration ability of colon cancer cells was detected by using cell counting kit-8 (CCK8) and Transwell method. The xenograft model of colon cancer in nude mice was constructed to observe the effect of miR-541-5p on tumor growth.Results:The relative expression level of miR-541-5p in colon cancer tissues was lower than that in pericarcinomatous normal tissues (0.45±0.06 vs. 1.00±0.12, t = 43.385, P < 0.01). The relative expression level of miR-541-5p was 0.46±0.03, 0.67±0.04, 0.57±0.06, 0.17±0.02, 1.00±0.15, respectively in colon cancer cell lines HT29, SW480, SW620, HCT116 and enterocyte line HIEC of the normal people, and the difference was statistiacally significant ( F = 5.621, P < 0.01); the relative expression level of miR-541-5p in all colon cancer cell lines was lower than that in enterocyte line HIEC of the normal people. HCT116 cells were selected to make the subsequent experiments. The predicted results of TargetScan showed that 3'UTR of CCND1 might have sites complementary to those of miR-541-5p. Dual luciferase reporter gene assay showed that CCND1 was the target gene of miR-541-5p, and miR-541-5p negatively regulated the expression of CCND1. CCK-8 method showed that cell proliferation rate of HCT116 was (2.00±0.16)%, (0.89±0.08)%, (2.56±0.23)%, respectively in miR-NC group, miR-541-5p group, miR-541-5p+CCND1 group, and the difference was statistically significant ( F = 6.715, P < 0.01); among HCT116 cells with the overexpression of miR-541-5p, the transfected CCND1 chould reverse the inhibitory effect of miR-541-5p on cell proliferation. Transwell results showed that the overexpression of miR-541-5p inhibited the cell migration ability of HCT116, while the co-transfection of miR-541-5p mimics and CCND1 could reverse the inhibitory effect. In the colon cancer nude mice xenograft model, the tumor mass and size of nude mice in miR-541-5p group was decreased compared with that in the control group (all P < 0.05). Conclusions:miR-541-5p inhibits cell proliferation and migration of colon cancer cells via negatively regulating CCND1, and inhibits tumor growth in xenograft model of colon cancer in nude mice, thereby acting as a tumor suppressor in colon cancer.

Objective To detect the long term effect of pure and multiple concussions on spatial cognitive of rats.Methods One hundred and eighty 7-week-old Spragne-Dawley male rats with weight of 280 ± 30g were chosen and randomly divided into a control group and a concussion group.The cerebral concussion was induced in the rats using a metallic pendulum striker concussive device.After the first strike,the brain injury group was randomly divided into a pure cerebral concussion(PCC)group and a multiple cerebral concussion(MCC) group.After the second strike,the MCC group was randomly divided into two-fold cerebral concussion(2MCC) group and three-fold cerebral concussion(3MCC) group.The striking interval was 24h.One,3 and 6 months after trauma,their cognitive function was tested using Morris water maze.Results One month later after injury,there was no significant difference in the escape latency between the control group and PCC group.Significant differences in the measurement were observed between the control/PCC group and 2MCC group on the 7th day after the injury,also between the control/PCC and 3MCC groups on the 6th and 7th day.And there were significant differences between the 2MCC and 3MCC groups on the 6th and 7th days.The non-platform test did not observe any significant differences among the four groups.Three months after injury,there was still no significant difference between the control group and PCC group,PCC and 2MCC groups,as well as 2MCC and 3MCC groups in the escape latency.However,there was significant difference between the control group and 2MCC group on the 5th,6th and 7th days,between the control group and 3MCC group on the 4th,5th,6th and 7th days,as well as between PCC group and 3MCC group on the 6th and 7th days.In the non-platform test,there was no significant difference between the control group and PCC group,between PCC group and 2MCC group,as well as between 2MCC group and 3MCC group.However,2MCC and 3MCC groups spent significantly less time in the former platform quadrant,when compared with the control group and 3MCC group spent significantly less time than PCC group.Six months after injury,significant differences in the escape latency were observed between the control group and PCC group on the 6th and 7th days,and 2MCC group on the 5th,6th and 7th days,also and 3MCC groups on the 2nd,3rd,4th,5th,6th and 7th days,still between PCC group and 2MCC group on the 6th and 7th days,as well as between PCC group and 3MCC group on the 4th,5th,6th and 7th days.Moreover,there was significant difference between 2MCC and 3MCC groups only on the 7th day.In the non-platform test,PCC group,2MCC group and 3MCC group spent significantly less time in the former platform quadrant compared with the control group.Moreover,in this test significant differences were found between PCC group and 2MCC/3MCC group,but not between 2MCC group and 3MCC group.Conclusion With the increase of cerebral concussion times,earlier and more serious damage of spatial cognition will appear,with a significant cumulative effect in rats.Such rat model can be used to study the pathological changes of cognitive impairment in chronic traumatic encephalopathy.