Objective·To analyze the root canal diameter and taper of mandibular first molars using cone-beam CT(CBCT),investigate age-related changes in these parameters,and propose optimized clinical protocols for root canal preparation.Methods·From October 2022 to October 2023,CBCT images of 240 healthy mandibular first molars(120 three-canal type cases and 120 four-canal type cases)were collected from patients aged 20?59 years at the Stomatology Center,Haikou Affiliated Hospital of Central South University Xiangya School of Medicine.The three-canal type and the four-canal type samples were respectively divided into four age groups(20?29,30?39,40?49,and 50?59 years),with 30 cases in each group.Root canal length was measured,and buccolingual and mesiodistal diameters were recorded at four levels(coronal,middle,apical,and foramen).The tapers of the coronal 1/3,middle 1/3,and apical 1/3 segments were calculated in both buccolingual and mesiodistal directions.Differences among the age groups were compared.Results·The buccolingual diameters of all root canals exceeded the corresponding mesiodistal diameters.At several levels of most root canals,the diameters in the<50-year-old groups were significantly larger than those in the 50?59-year-old group(P<0.05).The buccolingual tapers exceeded the corresponding mesiodistal tapers in all root canals of mandibular first molars.The mean mesiodistal tapers in different age groups ranged from 0.07 to 0.34 in the coronal 1/3 segment,0.03 to 0.09 in the middle 1/3 segment,and 0.05 to 0.11 in the apical 1/3 segment.At several levels of most root canals,the tapers in the<50-year-old groups were significantly larger than those in the 50?59-year-old group(P<0.05).Conclusion·Mandibular first molars exhibit age-related narrowing of root canal diameter and reduced taper in individuals aged≥50 years.For initial apical file selection,#15 files are recommended for the distal canals in the three-canal type,while#10 files are advised for all other canals in the three-canal type and all canals in the four-canal type.For patients aged 50 years or older,the files for the distal canals in the three-canal type can be adjusted to#10,and the files for the mesiobuccal and distallingual canals in the four-canal type to#8.Regarding preparation taper,a 0.08-taper orifice opener is recommended for the coronal 1/3 segment.During mid-lower segment preparation,0.04-taper master apical files are suggested for the mesiobuccal and mesiolingual canals in the three-canal type and the mesiolingual and distallingual canals in the four-canal type,0.06-taper files for the distal canals in the three-canal type and the mesiobuccal canals in the four-canal type,and 0.08-taper files for the distobuccal canals in the four-canal type.Root canal obturation is recommended to be performed using vertical compaction with large-taper gutta-percha cones combined with bioceramic sealer.

Objective To explore the therapeutic effect and mechanism of electroacupuncture on depressive model mice based on the phosphatase and tensin homolog-induced kinase 1(PINK1)/Parkin pathway.Methods Specific pathogen-free grade male C57BL/6J mice were used.For experiment 1,60 mice were randomly divided into blank,model,sham electroacupuncture,and electroacupuncture groups using the random number table method,with 15 rats per group.For experiment 2,30 mice were randomly divided into normal,cyclosporine A(CsA),and electroacupuncture+CsA groups using the same method,with 10 rats per group.The chronic restraint stress(CRS)was used to establish a depression model.After successful modeling,CRS was continued to maintain model stability.After modeling,1 h before daily CRS stimulation,the electroacupuncture and electroacupuncture+CsA groups received electroacupuncture interventions at the"Baihui"(GV20)and"Zusanli"(ST36)acupoints,using continuous wave stimulation at a frequency of 2 Hz and an intensity of 1 mA for 20 min,once daily for 7 consecutive days.Mice in the sham electroacupuncture group received superficial needling at non-meridian,non-acupoint locations under the axilla 1 h before CRS,with the electroacupuncture device connected but not powered on once a day for 7 consecutive days.Mice in the CsA and electroacupuncture+CsA groups received an intraperitoneal injection of CsA solution(0.2 mg/g)30 min before electroacupuncture intervention,once daily for 7 consecutive days.In experiment 1,depressive-like behavior was assessed using the open field,tail suspension,and sucrose preference tests.The spontaneous excitatory postsynaptic currents(sEPSC)and spontaneous inhibitory postsynaptic currents(sIPSC)parameters of hippocampal neurons were evaluated using brain slice patch clamp techniques.Western blotting was conducted to measure the expression levels of mitochondrial autophagy-related proteins PINK 1,phosphorylated PINK1(p-PINK1),Parkin,microtubule-associated protein 1A/1B-light chain 3-Ⅱ/Ⅰ(LC3-Ⅱ/Ⅰ),ubiquitin-binding protein(p62),and mitochondrial markers,including translocase of outer mitochondrial membrane 20(TOMM20),heat shock protein 60(HSP 60),and cytochrome c oxidase Ⅳ(COX Ⅳ).Immunofluorescence was used to detect PINK1 protein expression in the CA3 region of the hippocampus.Transmission electron microscopy was used to examine the ultrastructure of mitochondria in hippocampal neurons.On the basis of experiment 1,experiment 2 evaluated depressive-like behavior in mice using sucrose preference,open field,and tail suspension tests;Western blotting was used to detect the expressions of PINK1,p-PINK1,Parkin,LC3-Ⅱ/Ⅰ,p62,TOMM20,HSP 60,and COX Ⅳ proteins of hippocampus in mice.The mitochondrial ultrastructure was observed in hippocampal neurons using transmission electron microscopy.Results In experiment 1,compared with the blank group,the model and sham electroacupuncture groups exhibited a decrease in sucrose consumption rate,a decrease in the time spent in the center area,a reduced proportion of distance moved in the center area,and an increase in immobility time of tail suspension(P＜0.05).The sEPSC and sIPSC in hippocampal neurons decreased in both amplitude and frequency(P＜0.05).p-PINK1,Parkin,LC3-Ⅱ/Ⅰ,TOMM20,HSP 60,and COXⅣ expression levels in the hippocampus were reduced,whereas the p62 expression level was increased(P＜0.05).The average fluorescence intensity of PINK1 in the CA3 region of the hippocampus decreased(P＜0.05).The number of healthy mitochondria in hippocampal neurons was reduced,with numerous damaged mitochondrial structures observed.Compared to the model and sham electroacupuncture groups,the electroacupuncture group showed an increased in the time spent in the center area,a higher proportion of distance moved in the center area,and an elevated sucrose consumption rate.In contrast,the immobility time in the tail suspension test decreased(P＜0.05),whereas the amplitude and frequency of sEPSC and sIPSC in hippocampal neurons increased(P＜0.05).p-PINK1,Parkin,LC3-Ⅱ/Ⅰ,TOMM20,HSP 60,and COXⅣ expression levels in the hippocampus increased,whereas the p62 expression level decreased(P＜0.05).The average fluorescence intensity of PINK1 in the CA3 region of the hippocampus increased(P＜0.05).Additionally,mitochondrial damage in hippocampal neurons was alleviated,and a notable presence of autophagosomes mitophagy lysosomes was observed.In experiment 2,compared with the normal group,the mice in the CsA and electroacupuncture+CsA groups showed a decrease in the time spent in the center area and the proportion of distance moved in the center area,a decrease in sucrose consumption rate,and an increase in the immobility time in the tail suspension test(P＜0.05).p-PINK1,Parkin,LC3-Ⅱ/Ⅰ,TOMM20,HSP 60,and COX Ⅳ expression levels in the hippocampus decreased,whereas p62 expression increased(P＜0.05).Many damaged mitochondria were observed in hippocampal neurons.Conclusion Electroacupuncture may exert its antidepressant effects by promoting PINK1/Parkin pathway-mediated mitophagy to eliminate damaged mitochondria,thereby restoring the function of hippocampal neurons in depressive model mice.

Objective To explore the therapeutic effect and mechanism of electroacupuncture on depressive model mice based on the phosphatase and tensin homolog-induced kinase 1(PINK1)/Parkin pathway.Methods Specific pathogen-free grade male C57BL/6J mice were used.For experiment 1,60 mice were randomly divided into blank,model,sham electroacupuncture,and electroacupuncture groups using the random number table method,with 15 rats per group.For experiment 2,30 mice were randomly divided into normal,cyclosporine A(CsA),and electroacupuncture+CsA groups using the same method,with 10 rats per group.The chronic restraint stress(CRS)was used to establish a depression model.After successful modeling,CRS was continued to maintain model stability.After modeling,1 h before daily CRS stimulation,the electroacupuncture and electroacupuncture+CsA groups received electroacupuncture interventions at the"Baihui"(GV20)and"Zusanli"(ST36)acupoints,using continuous wave stimulation at a frequency of 2 Hz and an intensity of 1 mA for 20 min,once daily for 7 consecutive days.Mice in the sham electroacupuncture group received superficial needling at non-meridian,non-acupoint locations under the axilla 1 h before CRS,with the electroacupuncture device connected but not powered on once a day for 7 consecutive days.Mice in the CsA and electroacupuncture+CsA groups received an intraperitoneal injection of CsA solution(0.2 mg/g)30 min before electroacupuncture intervention,once daily for 7 consecutive days.In experiment 1,depressive-like behavior was assessed using the open field,tail suspension,and sucrose preference tests.The spontaneous excitatory postsynaptic currents(sEPSC)and spontaneous inhibitory postsynaptic currents(sIPSC)parameters of hippocampal neurons were evaluated using brain slice patch clamp techniques.Western blotting was conducted to measure the expression levels of mitochondrial autophagy-related proteins PINK 1,phosphorylated PINK1(p-PINK1),Parkin,microtubule-associated protein 1A/1B-light chain 3-Ⅱ/Ⅰ(LC3-Ⅱ/Ⅰ),ubiquitin-binding protein(p62),and mitochondrial markers,including translocase of outer mitochondrial membrane 20(TOMM20),heat shock protein 60(HSP 60),and cytochrome c oxidase Ⅳ(COX Ⅳ).Immunofluorescence was used to detect PINK1 protein expression in the CA3 region of the hippocampus.Transmission electron microscopy was used to examine the ultrastructure of mitochondria in hippocampal neurons.On the basis of experiment 1,experiment 2 evaluated depressive-like behavior in mice using sucrose preference,open field,and tail suspension tests;Western blotting was used to detect the expressions of PINK1,p-PINK1,Parkin,LC3-Ⅱ/Ⅰ,p62,TOMM20,HSP 60,and COX Ⅳ proteins of hippocampus in mice.The mitochondrial ultrastructure was observed in hippocampal neurons using transmission electron microscopy.Results In experiment 1,compared with the blank group,the model and sham electroacupuncture groups exhibited a decrease in sucrose consumption rate,a decrease in the time spent in the center area,a reduced proportion of distance moved in the center area,and an increase in immobility time of tail suspension(P＜0.05).The sEPSC and sIPSC in hippocampal neurons decreased in both amplitude and frequency(P＜0.05).p-PINK1,Parkin,LC3-Ⅱ/Ⅰ,TOMM20,HSP 60,and COXⅣ expression levels in the hippocampus were reduced,whereas the p62 expression level was increased(P＜0.05).The average fluorescence intensity of PINK1 in the CA3 region of the hippocampus decreased(P＜0.05).The number of healthy mitochondria in hippocampal neurons was reduced,with numerous damaged mitochondrial structures observed.Compared to the model and sham electroacupuncture groups,the electroacupuncture group showed an increased in the time spent in the center area,a higher proportion of distance moved in the center area,and an elevated sucrose consumption rate.In contrast,the immobility time in the tail suspension test decreased(P＜0.05),whereas the amplitude and frequency of sEPSC and sIPSC in hippocampal neurons increased(P＜0.05).p-PINK1,Parkin,LC3-Ⅱ/Ⅰ,TOMM20,HSP 60,and COXⅣ expression levels in the hippocampus increased,whereas the p62 expression level decreased(P＜0.05).The average fluorescence intensity of PINK1 in the CA3 region of the hippocampus increased(P＜0.05).Additionally,mitochondrial damage in hippocampal neurons was alleviated,and a notable presence of autophagosomes mitophagy lysosomes was observed.In experiment 2,compared with the normal group,the mice in the CsA and electroacupuncture+CsA groups showed a decrease in the time spent in the center area and the proportion of distance moved in the center area,a decrease in sucrose consumption rate,and an increase in the immobility time in the tail suspension test(P＜0.05).p-PINK1,Parkin,LC3-Ⅱ/Ⅰ,TOMM20,HSP 60,and COX Ⅳ expression levels in the hippocampus decreased,whereas p62 expression increased(P＜0.05).Many damaged mitochondria were observed in hippocampal neurons.Conclusion Electroacupuncture may exert its antidepressant effects by promoting PINK1/Parkin pathway-mediated mitophagy to eliminate damaged mitochondria,thereby restoring the function of hippocampal neurons in depressive model mice.

Objective·To analyze the root canal diameter and taper of mandibular first molars using cone-beam CT(CBCT),investigate age-related changes in these parameters,and propose optimized clinical protocols for root canal preparation.Methods·From October 2022 to October 2023,CBCT images of 240 healthy mandibular first molars(120 three-canal type cases and 120 four-canal type cases)were collected from patients aged 20?59 years at the Stomatology Center,Haikou Affiliated Hospital of Central South University Xiangya School of Medicine.The three-canal type and the four-canal type samples were respectively divided into four age groups(20?29,30?39,40?49,and 50?59 years),with 30 cases in each group.Root canal length was measured,and buccolingual and mesiodistal diameters were recorded at four levels(coronal,middle,apical,and foramen).The tapers of the coronal 1/3,middle 1/3,and apical 1/3 segments were calculated in both buccolingual and mesiodistal directions.Differences among the age groups were compared.Results·The buccolingual diameters of all root canals exceeded the corresponding mesiodistal diameters.At several levels of most root canals,the diameters in the<50-year-old groups were significantly larger than those in the 50?59-year-old group(P<0.05).The buccolingual tapers exceeded the corresponding mesiodistal tapers in all root canals of mandibular first molars.The mean mesiodistal tapers in different age groups ranged from 0.07 to 0.34 in the coronal 1/3 segment,0.03 to 0.09 in the middle 1/3 segment,and 0.05 to 0.11 in the apical 1/3 segment.At several levels of most root canals,the tapers in the<50-year-old groups were significantly larger than those in the 50?59-year-old group(P<0.05).Conclusion·Mandibular first molars exhibit age-related narrowing of root canal diameter and reduced taper in individuals aged≥50 years.For initial apical file selection,#15 files are recommended for the distal canals in the three-canal type,while#10 files are advised for all other canals in the three-canal type and all canals in the four-canal type.For patients aged 50 years or older,the files for the distal canals in the three-canal type can be adjusted to#10,and the files for the mesiobuccal and distallingual canals in the four-canal type to#8.Regarding preparation taper,a 0.08-taper orifice opener is recommended for the coronal 1/3 segment.During mid-lower segment preparation,0.04-taper master apical files are suggested for the mesiobuccal and mesiolingual canals in the three-canal type and the mesiolingual and distallingual canals in the four-canal type,0.06-taper files for the distal canals in the three-canal type and the mesiobuccal canals in the four-canal type,and 0.08-taper files for the distobuccal canals in the four-canal type.Root canal obturation is recommended to be performed using vertical compaction with large-taper gutta-percha cones combined with bioceramic sealer.

Objective To investigate the features and characteristics of GLIS3 gene mutation in patients with congenital hypothyroidism(CH),and to establish the theoretical basis for gene diagnosis and prenatal diagnosis of CH.Methods Genomic DNA was extracted from peripheral blood leukocytes of 50 patients with CH who were collected from February 2007 to November 2016 in Shandong Province.The exon 2 to 11 of GLIS3 were amplified with 11 pairs of sequence specific primers designed by Primer 5.0.Polymerase chain reaction and the first generation of sequencing method(Sanger sequencing) were used to detect the mutation.Comparison of the sequencing results with the GLIS3 reference sequence (National Center for Biotechnology Information Reference Sequence:NC_000009.12) helped to screen gene mutations.Results The 50 CH patients included 22 boys and 28 girls,and the sex ratio was 1.0 ∶ 1.3.The mean age was (2.5 ± 0.5) years.Six cases (12％) had thyroid gland hypoplasia,23 cases (46％) had thyroid gland agenesis and 21 cases(42％) with ectopic thyroid gland.C2507A missense mutation was found in exon 10 of GLIS3 in a thyroid gland agenesis case,which might result in proline to glutamine substitution at codon 836.One mutant (rs780019691,c.C289T) was detected which was nonsense mutation (Arg→Stop) in another thyroid gland agenesis child.Conclusions The mutation rate of GLIS3 gene is very low in CH children of Shandong province.Further studies are needed to investigate the relationship between GLIS3 genotypes and clinical phenotypes.

Objective To study the expression and the mechanism of miR-155in the villi of patients with unexplained recurrent spontaneous abortion (URSA).Methods The expression of miR-155 in the villi of 36 cases with URSA (URSA group) and 25 women with normal early pregnancy (control group) were detected by stem-loop real-time reverse transcription (RT) qPCR.Expression of hypoxia inducible factor-1 (HIF-1α),vascular endothelial cell growth factor(VEGF) and micro lymphatic vessel density (MVD) in the villi of were measured by immnohistochemical staining among two groups.Results (1) miR-155 expression:the mean miR-155 expression were 1.456 (0.489,2.459) in URSA group and 2.833 (1.740,3.794) in control group,which reached statistical difference (P ＜0.05).The mean expression of miR-155 of 1.683 (0.902,2.459) in URSA group with abortion times (≤ 3) was significantly higher than 1.229 (0.489,1.719) in URSA group with more than 4 times abortion (P ＜ 0.05).(2) Indexes:the expression of HIF-1α,VEGF and MVD value were 121 ± 12,134 ± 12,36 ± 6 in URSA group and 99 ± 10,109 ± 10,28 ±4 in control group,which reached statistical difference(P ＜ 0.01).The expression of HIF-1α,VEGF and MVD value of 119 ± 12,134 ± 12,35 ± 5 in URSA group with less than 3 times abortion was significantly lower than 128 ± 12,138 ± 12,43 ± 6 in URSA group with more than 4 times abortion (P ＜ 0.01).Conclusions The expression of miR-155 and HIF-1α is topically stimulated by oxygen signal.HIF-1α adjusts the transcription and translation of VEGF,which together involved in placental trophoblast invasion and placental angiogenesis.The low expression of miR-155 could interfere with expression of HIF-1α and VEGF,which might be involved in villous vascular dysplasia in URSA.