Objective To study the validity of acoustic voice quality index（AVQI） in Mandarin Chinese language, to investigate the accuracy of AVQI in discriminating normal and dysphonic voice, and the accuracy in discriminating across the perceptual categories of dysphonia severity. Methods This study included 59 subjects with normal voice and 217 subjects with dysphonic. Voice samples of sustained vowel/a/and continuous speech were collected. AVQI was calculated using AVQI（v02.06） script in Praat software. G score of GRBAS system was assessed by 5 raters and the mean value was used in following analysis. Spearman correlation coefficient was calculated. ROC curves were drawn to evaluate the accuracy of discriminating normal and dysphonic voice, and discriminating across the perceptual categories of dysphonia severity. Results The correlation coefficient of AVQI and Gmean was 0.756（P＜0.001）. When discriminating normal and dysphonic voice at Gmean=0.2, AUC=0.849 and threshold was 2.905. When discriminating them under clinical diagnosis, AUC was 0.788 and threshold was 3.18. When discriminating voice of different severity, AUC and threshold of normal vs slight were 0.770 and 1.985 respectively. Those of slight vs moderate were 0.755 and 4.17, and those of moderate vs severe were 0.799 and 6.36 respectively. Conclusion The AVQI is an easy-to-use and quick tool for the objective measure of overall voice quality in Mandarin Chinese language. It could discriminate normal and dysphonic voice, and the voice with different dysphonia severity to a certain extent.

目的 分析未经嗓音训练或改变音调手术治疗的跨性别女性的嗓音特点,探讨其与顺性别人群的差异.方法 选择78例未经嗓音训练或改变音调手术的跨性别女性为研究对象,以顺性别男性和女性各30例为对照组,分别采集汉语习惯性言语样本和发元音/a/、/i/声样本,分析言语平均基频(SFF)、最高言语基频(H-SFF)、最低言语基频(L-SFF),元音/a/的最高基频、最低基频、jitter、shimmer、最长发声时间(M PT)以及元音/a/、/i/的第1、2、3共振峰(F1、F2、F3)频率.另招募男性、女性志愿者各10例,对所有受试者的嗓音进行女性化程度主观听觉评估(1～2分为女性化嗓音,3～5分为中性化嗓音,6～7分为男性化嗓音).结果 跨性别女性的SFF、H-SFF均显著高于男性对照组(P<0.01),低于女性对照组(P<0.01),但音域范围与男性对照组无明显差异(P>0.05);跨性别女性组MPT显著小于男性对照组(P<0.01),与女性对照组差异无统计学意义(P>0.05);跨性别女性组的元音/a/、/i/的F2、F3频率显著高于男性对照组(P<0.01),元音/a/的F2、元音/i/的F1低于女性对照组(P<0.05).跨性别女性组的嗓音女性化程度评分(3.397±1.36分)介于男性(6.138±0.58分)和女性对照组(1.75±0.61分)之间.结论 本组跨性别女性的嗓音特征介于顺性别男性与女性之间,未经嗓音训练或改变音调手术治疗的跨性别女性难以达到女性的嗓音条件和性别认同,该类群体可能存在改变嗓音的诉求.

子宫肌瘤是女性生殖系统最常见肿瘤,严重危害育龄期妇女的身心健康.近年来,随着肿瘤免疫学的快速发展和中医药研究的不断深入,有关中药免疫调控抗肿瘤的药物机制陆续被发现.中医正气学说与免疫系统具有相似之处,中药多靶点、多通路、双向调节的特点在免疫调控机制中具有突出优势.本文从调控T淋巴细胞、自然杀伤细胞、树突状细胞、调节性T细胞和免疫检查点5个方面对子宫肌瘤治疗中使用频率较高的中药或复方涉及的免疫分子机制进行阐述,旨在为更好地开展子宫肌瘤中医药治疗提供理论支持.

子宫肌瘤是一种常见的良性妇科肿瘤,多发生在育龄期女性,与雌激素浓度密切相关[1-3].本病发病率高,大多数患者临床症状不明显,一般随着年龄增长,雌孕激素浓度的下降,可自行萎缩,但有部分患者子宫肌瘤过大或者生长过快,症状明显可能出现经血增多及其引起的贫血,过大的子宫肌瘤产生压迫症状如便秘、尿潴留、腰痛,甚至流产,影响受孕,严重影响女性的正常生活[4-7] ,极少数患者向恶性肿瘤转化[8].目前临床上尚缺乏有效的药物治疗,一般单发性肌瘤超过 5 cm 或者多发出现明显症状者建议手术治疗[9-13].

Objective To assess true vocal fold (TVF) length and cricothyroid distance(CTD) with ultrasonography in male-to-female (MtF)transsexual voice surgery.Methods Five MtF transsexuals were divided into two groups according to their voice change surgery methods.High-frequency ultrasonography was used to measure the length of true vocal folds and cricothyroid distance.Measurements were compared pre-and post-surgery, and correlations with fundamental frequency (F0) were analyzed.Results The ultrasonography measurements clearly showed the laryngeal structure and TVF measurement marks.After vocal fold shortening and retrodisplacement of anterior commissure(VFSRAC) surgery, the ultrasonographic measurements showed that the shortening length of TVF were 0.37 cm(21%),0.69 cm(37%) and 0.40 cm(25%), respectively.The CTD in ultrasonographicimages were 1.0 cm pre-surgery of cricothyroid approximation (CTA) and-0.33 post-surgery of CTA.The displacement of cricoid cartilage was 1.33 cm, which was consistent with the laryngeal CT image.Conclusion Both TVF length and CTD can be shown by high-frequency ultrasonography, which can be used to quantitatively assessment and follow-up MtF transsexual voice surgery.

Objective To explore the feasibility and practicability of research electronic data capture (REDCap) database in clinical research of phonosurgery.Methods REDCap database was designed and constructed to collect the basic information,personal history and history of professional voice use,related diseases,physical examination,voice assessment,voice handicap index(Simplified Chinese version),surgical information,pathological result,reflux symptom index,reflux finding score (RFS).For each patient,6 data collection time-points were set,including enrollment,operation,one week,3 weeks,2 months and one year after operation.The application of REDCap database in our hospital from July 2016 to Jan.2017 was summarized.Results Consisted of 10 instrument tables,3 modules and 6 event timepoints,our voice REDCap database was able to achieve simultaneous multiple-ended input of medical data,quality control and authority management.104 patients were enrolled in our REDCap database.Of them,102 completed their follow up of one-week and three-week,and 98 completed two-month follow up with all instruments complished.Conclusion With advantages of integrity and timeliness of record,decreased lost rate of follow up,our REDCap database can cover the basic flow and meet demands of clinical research of phonosurgery.

目的 探讨针对青春期假声发音模式的综合嗓音矫治方法的有效性.方法 回顾性分析12例患者的嗓音评估状况,采用针对性综合嗓音矫治方案后,对比分析嗓音矫治前后的基频、基频声学参数、语图、喉的位置、呼吸方式及声门闭合情况.结果 12例患者均存在喉位置高、胸式呼吸、声门关闭不严等发音模式,综合嗓音矫治后,患者基频F0明显降低,基频标准差F0sd明显减轻,最大发音时无明显差异,呼吸方式转变为腹式或胸腹式,喉位置发音时稳定.结论 针对青春期假声发音模式的综合嗓音矫治方法疗效明显.

Objective To investigate the value of clinical characteristics in diagnosis of vocal fold paralysis (VFP) and arytenoid dislocation .Methods Eighty - eight patients of VFP and 27 patients of arytenoid dislocation were studied , by comparing the causes , laryngeal morphologic characteristics and laryngeal electromyography (LEMG) .Results The causes of 88 VFP patients included surgery (45 cases) ,neck trauma(2 cases) ,idiopathic causes(16 cases) ,infection(16 cases) ,and tumor invasion - related(9 cases) .Of the 27 arytenoid dislocation pa‐tients ,24 had a history of endotracheal intubation and the others had a history of gastric tube insertion .The vocal folds were mostly fixed at the paramedian position ,followed by the abducent position and the median position .No significant differences were found in laryngeal morphologic characteristics between the two groups ,including vocal fold shape , glottis vertical symmetry , mucosal waves , supraglottic compensation , glottis closure and arytenoid movement .The LEMG of VFP patients appeared as denervation patterns ,reinnervation potentials ,or electrical si‐lence ;the recruitment patterns appeared as mix or simple patterns ;the evoked potentials were absent .Of the VFP patients ,54 cases(61 .36% % )were found synkinesis of involved posterior cricoarytenoid and two of them also in ‐volved thyroatenoid .The patients with synkinesis had lower percentage of vocal fold bowing and higher percentage of glottic vertical asymmetry compared to the ones without synkinesis .Of the VFP patients whose cause was surgery or neck trauma ,the median - position fixed vocal folds were mostly observed in the patients with duration of less than 1 month or with synkinesis .Of the 27 arytenoid dislocation patients ,20(74 .07% )showed normal LEMG pat‐terns and 7(25 .93% )showed apparent LEMG abnormality on the affected side .Conclusion The causes of vocal fold paralysis and arytenoid dislocation are different .Laryngeal morphologic characteristics have limitations in distinguis‐hing vocal fold paralysis from arytenoid dislocation .The shape and position of involved vocal folds of the VFP pa‐tients are correlated with duration ,nerve regeneration and synkinesis .

Objective To evaluate the role of serum GP73 for diagnosing decompensated cirrhosis in population with chronic HBV infections.Methods The present study included three populations:200 patients with chronic HBV infections and receiving liver biopsy; 200 patients with decompensated cirrhosis;and 200 patients with hepatocellular carcinoma.All patients were HBsAg-positive over six months.Results Comparing with those in the patients with chronic hepatitis B (67.38 ±57.45 ng/ml),the serum GP73 levels in cirrhosis patients (221.9 ± 108.5 ng/ml) and HCC patients (152.4 ± 102.7 ng/ml) were significantly increased.Taken the population with chronic hepatitis B as "control group",the patients with decompensated cirrhosis as "patients group",the area of ROC analysis was 0.91 (95％ CI:0.88-0.94) (P ＜0.0001).Set the cut-off value at 150 ng/ml,the diagnosing sensitivity and specificity were 72.5％ and 93.5％,respectively.Conclusion For patients with chronic HBV infections and a higher GP73 levels,decompensated cirrhosis should be considered,except diagnosis of hepatocellular carcinoma.

Objective To construct the prokaryotic expression vector p ET-32a（+）-C2orf69 and induce the expression of recombinant proteins in vitro. Then the possible effects of recombinant protein on cell proliferation was observed and rabbit-anti-C2orf69 protein polyclonal antibodies was obtained.Methods Gene fragment of C2orf69 was amplified by PCR and then prokaryotic expression plasmid pE T-32a（+）-C2orf69 was constructed. Recombinant protein C2orf69 expression was identified by SDS-PAGE and Western blot. The white-ear rabbits were immunized with purified recombinant protein C2orf69, and the potency and specificity of polyclonal antibody were evaluated by enzyme-linked immunosorbent assay（ELISA） and Western blot. Also, different liver cells were incubated with recombinant protein C2orf69 in vitro. Results C2orf69 gene fragment was successfully amplified, results of gene sequencing were consistent with the sequence in Gen Bank. Recombinant protein of C2orf69 was successfully induced and expressed. The polyclonal antibody titer was up to 1︰1 280 000 through enzyme-linked immunosorbent assay. Results of cell proliferation showed that the recombinant protein could inhibit the proliferation of different liver cells. Conclusions The recombinant protein C2orf69 could inhibit the proliferation of different liver cells, and we speculated that it may be a widely roled inhibitor of hepatocyte proliferation. Our experiment showed that the proliferation inhibition of cells may be realized by G1 phase extending and S phase shortening.

Objective To compare the advantages and disadvantages of the three isolation regents for isolating primary hepatic stellate cells(HSCs) of mice.Methods The mice HSCs was isolated by in situ perfusion of per-perfusion solution,perfusion with collagenase Ⅳ.After digestion,Ficoll-Paque PLUS,perco11 and Nycodenz were use for the gradient centrifugation.To calculate the quantity,purity and the viability of the HSCs.The autofluorescence and Oil O staining were used to identify primary HSCs.Results The quantity of HSCs was (1.0 ± 0.5) × 106,(1.0 ± 0.5) × 105,(0.8 ± 0.5) × 106/ g_liver,respectively.Using Ficoll-Paque PLUS,percoll and Nycodenz,the purity of primary HSCs were 75％-85％,91％-95％,91％-95％,respectively.The viability were all above 90％.Conclusion compared with Ficoll-Paque PLUS and percoll,the Nycodenz is the the best separating medium for isolation and purification of mice primary HSCs.

Objective To prepare an Colgalt2 ELISA kit using Colgalt2 N-terminal and C-terminal polyclonal antibody.Methods Colgalt2 partial CDs were amplified from HepG2 cell,and inserted into prokaryotic expression plasmids,and induced the expression of N-terminal recombinant Colgalt2 protein.N-terminal Colgalt2 polyclonal antibody was prepared by immunizing rabbits with purified recombinant protein.Titer and specificity of the antibody was determined by ELISA and Western Blot.Microtiter plate was coated with N-terminal Colgalt2 polyclonal antibody,and other polyclonal antibody of Colgalt2 C-terminal was labeled by HRP.Results We successfully cloned N-terminal of Colgalt2 and obtained the recombinant Colgalt2 protein.ELISA analysis showed that titer of anti-Colgalt2 was higher than 1 ∶ 1 280 000.Western Blot analysis showed that the antibody with a good specificity.We successfully developed an ELISA kit of Colgalt2.The data of the different concentrations of recombinant protein Colgalt2 showed that the ELISA kit had a good sensitivity.From1.25 μg/ml to 25μg/ml,the coefficient of linear correlation was 0.98.Conclusion The Colgalt2-ELISA kit may be used to determine Colgalt2 levels in samples.

Although Golgi protein 73 (GP73) has been widely evaluated for diagnosing hepatocellular carcinoma (HCC) and other liver diseases in recent decade, its serum profile of patients with hepatitis B virus (HBV)-associated acute-on-chronic liver failure (HBV-ACLF) is still unknown. This study was designed to evaluate the serum levels of GP73 in patients with HBV-ACLF. The participants included 200 apparently healthy controls; 200 patients with chronic hepatitis B (CHB); 200 patients with HCC; 210 patients with HBV-ACLF, in which 29 HBV-ACLF patients were followed up for 3 months. All patients were Hepatitis B virus surface antigen (HBsAg) positive. The concentrations of GP73 in patients with HBV-ACLF (285.3 ± 128.5 ng/mL) were markedly higher than those HCC patients (159.1 ± 105.8 ng/mL), CHB patients (64.65 ± 44.99 ng/mL), and healthy controls (35.37 ± 12.41 ng/mL). When the cut-off value was set at 182.1 ng/mL, the sensitivity and specificity of HBV-ACLF diagnosis were 77.62% (95% confidence interval [CI]: 71.37%-83.07%) and 95.50% (95% CI: 92.27%-98.26%), respectively. If serum GP73 concentration was still above 361.6 ng/mL after 14 days of follow-up, the patient's prognosis may be depressed. Serum GP73 may be used to diagnosis HBV-ACLF in population with chronic HBV infections.

Objective To create mouse model of glycogene GLT25D2 knockout,for exploring the molecular mechanism of collagen glycosylation.Methods The neomycin cassette was served as the positive selection marker during the embryonic stem cells targeting step,it was removed by expressing Cre recombinase via a plasmid in the targeted ES cells.Partial exons 2 and total exons 3 were removed,and replaced with loxpNeoloxp.The translation was stopped in exons 2 when meeting the stop code,which is added in-framely into exons 2.The PCR technique was used to identify genotype.Results After stem cells targeting,four founder mice were produced (2 female and 2 male).Total 163 mice were reproduced,including 40 GLT25D2-/-mice,89 GLT25D2+/-mice,and 34 GLT25D2+/+ mice,after eight months breeding.The proportion was consistent with the Mendelian genetic law.But the GLT25D2-/-mice showed a relatively low reproductive capability.The GLT25D2-/-mice birth rate of each embryo is 3-4,less than those of GLT25D2 +/-mice (each embro 6-8).More importantly,the average weigh of GLT25D2-/-mouse significantly higher than those of GLT25D2 +/-mice,and GLT25D2 +/+ mice,at 20,40,and 60 days after birth.The variation trend was gradually decreased with the mice growth.Conclusion The GLT25D2 knockout mice showed a relatively low reproductive capability and abnormal development.Compared with the GLT25D2 +/+ mice,the weight of GLT25D2-/-mice was increased.

OBJECTIVE:To explore the levels of serum GP73 in patients with fatty liver disease.METHODS:The sera GP73 were determined by ELISA in 178 patients with fatty liver disease and 100 healthy controls.RESULTS:Serum GP73 levels were significantly increased in patients with various fatty liver diseases(70.62 +/- 60.60 ng/ml), compared with those of control population (35.61 +/- 12.22 ng/ml). In patients with alcoholic fatty liver disease, acute liver injury, chronic hepatitis B, and non-alcoholic fatty liver disease, their serum GP73 concentration were 81.86 +/- 47.82 ng/ml, 82.77 +/- 77.73 ng/ml, 63.84 +/- 50.62 ng/ml, and 65.75 +/- 62.20 ng/ml, respectively. But no significant difference was found between these groups (P > 0.05). In 68 patients with F > or = 1.0 (71.46 +/- 66.48 ng/ml), 75 patients with F> or = 2.0 (69.58 +/- 62.31 ng/ml), and 34 patients with F3-F4 (71.65 +/- 43.89 ng/ml), there were also no marked differences was observed between these fatty groups (F = 0.02, P = 0.98).CONCLUSION:Serum GP73 levels were increased in patients with different liver diseases, but its concentrations were seems not related with degree of fatty injury.

目的 初步探讨蛋白质O-糖基化抑制剂Benzyl-α-GalNAc对肝细胞的影响.方法 取雌性Balb/c小鼠(8～10周)20只随机分为正常组(n =10),Benzyl-α-GalNAc药物组(n=10),给Benzyl-α-GalNAc药物(5 mg/kg,1次/d)灌胃.第2周末取小鼠血清及肝脏进行血清生化指标ALT及ALP测定及肝组织HE染色.培养HepG2细胞,Benzyl-α-GalNAc分别以0.5、1 及5 mg/ml与HepG2细胞共孵育.24 h后进行油红O染液染色.5 mg/ml药物组完全未贴壁,去除处理因素,继续培养12 h观察细胞.Benzyl-α-GalNAc以3 mg/ml处理HepG2细胞12 h及24 h观察细胞贴壁情况.培养L02细胞,Benzyl-α-GalNAc分别以0.1及0.5 mg/ml与L02细胞共孵育.24 h后进行油红O染液染色.结果 药物组与正常对照组比较,肝组织切片显示轻微脂肪变.油红O染色显示药物处理组HepG2及L02细胞内的脂滴积聚.高浓度组细胞完全没有贴壁,24 h后更换无药物培养基,去除处理因素12 h后,可见HepG2开始贴壁生长.结论 高浓度O-糖基化抑制剂Benzyl-α-GalNAc抑制体外肝细胞的黏附,并诱导轻微肝细胞脂肪变.

目的 体外克隆表达THEM6,诱导重组蛋白,制备多克隆抗体,明确在不同组织细胞的表达特征.方法 以外周血单核细胞总RNA反转录得到的cDNA为模板,扩增THEM6基因片段,诱导剂IPTG诱导THEM6重组蛋白表达,制备多克隆抗体,分析特异性、检测效价.用CCK-8检测试剂盒,分析THEM6重组蛋白对Jurkat细胞增殖的影响;观察该蛋白的组织、细胞系表达分布.结果 成功体外克隆THEM6,获得重组蛋白及抗THEM6的多克隆抗体,ELISA(enzyme-linked immuno sorbent assay)酶联免疫吸附测定检测效价>1 280 000;低浓度THEM6重组蛋白对Jurkat细胞增殖有影响,THEM6蛋白在Jurkat细胞等多种细胞及淋巴、肝脏等多种组织中表达.结论 THEM6基因在多种组织细胞中有广泛表达.

Objective To prepare the polyclonal antibody of C12orf28 and observe the expression features of C12orf28 in immunologic tissues. Methods The peptide sequence with high immunogenicity and specificity that contains 257 amino acids of the carboxy-terminal was selected and amplified by reverse transcription PCR. The DNA fragment C12orf28C257 was inserted into pET32a(+) plasmid and transformed into Escherichia coli BL21. The protein expression was induced with IPTG in vitro. Anti-C12orf28 was prepared by immunizing the big ears rabbit and was analyzed by Western blot and ELISA. The distribution of C12orf28 in tissues was detected by Western blot. Results The DNA fragment C12orf28C257 was cloned and its recombinant protein was successfully induced. The antibody titer was more than 1︰1.28 × 106 by ELISA and the high specificity of the polyclonal antibody was detected by Western blot. Conclusion The new gene C12orf28 is expressed in the fetal tissues such as intestine, lymph node, thymus and myocardiun, etc.

Objective To discuss the relationship between serum concentration of glycosyltransferase 25 domain 1(GLT25D1) and the grading of fibrosis in patients with chronic HBV infection.Methods The GLT25D1 gene was divided into two parts of GLT25D1-1 and GLT25D1-2 by antigenic epitope prediction software BepiPred 1.0 Server.With reverse transcription polymerase chain reaction(RT-PCR),genes(GLT25D1-1 and GLT25D1-2) were cloned into prokaryotic expression vector pET-32a(+).The potency and specificity of polyclonal antibody were evaluated by enzyme-linked immunosorbent assay(ELISA) and Western blot.An antigen to GLT25D1 ELISA double antibody sandwich method was used to detect,the serum of GLT25D1 in patients with various stages in the process of HBV infection of the liver.Results ELISA analysis indicated that the titer of the polyclonal antibody can be achieved 1∶ 1 280 000.The high specificity of the polyclonal antibody was confirmed by Western blot.To detect the serum of HBV infection at different stages by ELISA,we found that there were statistically significant differences of GLT25D1 OD values in chronic hepatitis B(≤S3),cirrhosis(S3-4 S4) and liver cancer patients compared with the normal group(P0.01).Conclusion With the serological testing results of chronic hepatitis,cirrhosis,and liver cancer patients,we found GTL25D1 exist in the process of HBV infection,and thus we speculated serum GLT25D1 detection can be used as an objective indicator to detect liver injury,and to assist other detection indicators better to provide the impetus for the diagnosis of hepatitis HBV infection process.