1.Biological function of tRNA-derived small RNA and its expression and clinical significance in liver diseases
Yinli LI ; Yan XU ; Zhiwei GUAN ; Lu MENG ; Yitong QU ; Jianli QIU
Journal of Clinical Hepatology 2025;41(6):1227-1234
Liver diseases cannot be easily detected in the early stage, and although invasive diagnostic methods, such as liver biopsy, are relatively accurate, they tend to have a low degree of acceptance, which greatly limits the improvement in diagnosis and treatment techniques for liver diseases. Therefore, it is of great importance to search for new biomarkers and therapeutic targets. As an emerging biomarker for liquid biopsy, tRNA-derived small RNA (tsRNA) is abnormally expressed in various liver diseases including viral hepatitis, fatty liver disease, liver injury, and liver cancer, and it can affect the development and progression of liver diseases by regulating the biological functions such as gene expression, epigenetic regulation, and protein translation. This article reviews the origin, classification, and biological function of tsRNA, as well as the research advances in tsRNA as biomarkers and potential therapeutic targets for liver diseases, so as to provide ideas for the early diagnosis and treatment of liver diseases.
2.Predict the effect of the number of positive preoperative serum tumor markers on the surgical method and prognosis of intrahepatic cholangiocarcinoma patients based on mediation analysis
Zonglong LI ; Jialu CHEN ; Yue TANG ; Delong QIN ; Chen CHEN ; Yinghe QIU ; Hong WU ; Yu HE ; Xianhai MAO ; Wenlong ZHAI ; Jingdong LI ; Xiao LIANG ; Chuandong SUN ; Kai MA ; Zhimin GENG ; Zhaohui TANG ; Zhiwei QUAN
Chinese Journal of Surgery 2024;62(7):685-696
Objective:To investigate the effect of the number of positive preoperative serological tumor markers on the surgical approach and prognosis of patients with intrahepatic cholangiocarcinoma.Methods:This is a retrospective case-series study. Data from 548 patients with intrahepatic cholangiocarcinoma after radical resection from October 2010 to April 2019 were retrospectively collected in 10 hospitals of China. There were 277 males and 271 females with an age of (57.8±10.2)years(range:23 to 84 years). Four hundred and twenty-six patients(77.7%) had at least one positive preoperative serum tumor marker. The data collection included the results of 4 preoperative serological tumor markers,other preoperative indicators(5 prodromal symptoms, 6 medical history,8 preoperative serological indicators,5 preoperative imaging indicators,and 14 preoperative pathological examination indicators),baseline data (gender and age),surgical methods,and prognostic follow-up data. Four preoperative results of serologic tumor marker and surgical procedure were converted into categorical variables. The number of positive preoperative serum tumor markers was used as the treatment variable,the surgical method was used as the mediating variable,and the survival time was used as the outcome variable. Univariate and multivariate analysis were used to screen for other preoperative indicators which were independent factors that influenced the surgical procedure and the prognosis of patients as covariates to analyze the mediating effect.Results:Of the 548 patients included in the study, 176 patients (32.1%) underwent partial hepatectomy,151 patients(27.5%) underwent hemihepatectomy, and 221 patients(40.3%) underwent partial hepatectomy or hemihepatectomy combined with other treatments. The results of the univariate and multivariate analysis showed that the number of positive serum tumor markers,intrahepatic bile duct dilatation,portal vein invasion,pathological differentiation,pathological type,vascular invasion,T stage,N stage and maximum tumor diameter were independent factors influencing the surgical procedure(all P<0.05). Intrahepatic bile duct dilatation,pathological differentiation and T stage were independent prognostic factors for patients with intrahepatic cholangiocarcinoma(all P<0.05). Intrahepatic bile duct dilatation,differentiation and T stage were included as covariates in the mediation effect model. The results showed that the number of positive serum tumor markers before surgery had a negative predictive effect on the survival time of patients with intrahepatic cholangiocarcinoma ( β=-0.092, P=0.039),and had a positive predictive effect on the surgical method ( β=0.244, P<0.01). The number of positive serum tumor markers had a negative predictive effect on the survival time of patients with intrahepatic cholangiocarcinoma ( β=-0.151, P=0.002). Direct and indirect effects accounted for 71.3% and 28.7% of total effects,respectively. Conclusions:The higher the positive number of preoperative tumor markers,the worse the prognosis of patients with intrahepatic cholangiocarcinoma. The number of positive cells not only directly affects the prognosis of patients,but also indirectly affects the prognosis of patients by affecting the surgical method.
3.Predict the effect of the number of positive preoperative serum tumor markers on the surgical method and prognosis of intrahepatic cholangiocarcinoma patients based on mediation analysis
Zonglong LI ; Jialu CHEN ; Yue TANG ; Delong QIN ; Chen CHEN ; Yinghe QIU ; Hong WU ; Yu HE ; Xianhai MAO ; Wenlong ZHAI ; Jingdong LI ; Xiao LIANG ; Chuandong SUN ; Kai MA ; Zhimin GENG ; Zhaohui TANG ; Zhiwei QUAN
Chinese Journal of Surgery 2024;62(7):685-696
Objective:To investigate the effect of the number of positive preoperative serological tumor markers on the surgical approach and prognosis of patients with intrahepatic cholangiocarcinoma.Methods:This is a retrospective case-series study. Data from 548 patients with intrahepatic cholangiocarcinoma after radical resection from October 2010 to April 2019 were retrospectively collected in 10 hospitals of China. There were 277 males and 271 females with an age of (57.8±10.2)years(range:23 to 84 years). Four hundred and twenty-six patients(77.7%) had at least one positive preoperative serum tumor marker. The data collection included the results of 4 preoperative serological tumor markers,other preoperative indicators(5 prodromal symptoms, 6 medical history,8 preoperative serological indicators,5 preoperative imaging indicators,and 14 preoperative pathological examination indicators),baseline data (gender and age),surgical methods,and prognostic follow-up data. Four preoperative results of serologic tumor marker and surgical procedure were converted into categorical variables. The number of positive preoperative serum tumor markers was used as the treatment variable,the surgical method was used as the mediating variable,and the survival time was used as the outcome variable. Univariate and multivariate analysis were used to screen for other preoperative indicators which were independent factors that influenced the surgical procedure and the prognosis of patients as covariates to analyze the mediating effect.Results:Of the 548 patients included in the study, 176 patients (32.1%) underwent partial hepatectomy,151 patients(27.5%) underwent hemihepatectomy, and 221 patients(40.3%) underwent partial hepatectomy or hemihepatectomy combined with other treatments. The results of the univariate and multivariate analysis showed that the number of positive serum tumor markers,intrahepatic bile duct dilatation,portal vein invasion,pathological differentiation,pathological type,vascular invasion,T stage,N stage and maximum tumor diameter were independent factors influencing the surgical procedure(all P<0.05). Intrahepatic bile duct dilatation,pathological differentiation and T stage were independent prognostic factors for patients with intrahepatic cholangiocarcinoma(all P<0.05). Intrahepatic bile duct dilatation,differentiation and T stage were included as covariates in the mediation effect model. The results showed that the number of positive serum tumor markers before surgery had a negative predictive effect on the survival time of patients with intrahepatic cholangiocarcinoma ( β=-0.092, P=0.039),and had a positive predictive effect on the surgical method ( β=0.244, P<0.01). The number of positive serum tumor markers had a negative predictive effect on the survival time of patients with intrahepatic cholangiocarcinoma ( β=-0.151, P=0.002). Direct and indirect effects accounted for 71.3% and 28.7% of total effects,respectively. Conclusions:The higher the positive number of preoperative tumor markers,the worse the prognosis of patients with intrahepatic cholangiocarcinoma. The number of positive cells not only directly affects the prognosis of patients,but also indirectly affects the prognosis of patients by affecting the surgical method.
4.Efficacy of different laparoscopic surgeries for gastrointestinal stromal tumors of gastric cardia and fundus: a multicenter study
Weifu ZHANG ; Xingyu FENG ; Peng ZHANG ; Wenjun XIONG ; Zaisheng YE ; Tao CHEN ; Haibo QIU ; Yuesheng YANG ; Wei WANG ; Luchuan CHEN ; Jiang YU ; Junjiang WANG ; Deqing WU ; Zhiwei ZHOU ; Kaixiong TAO ; Yong LI
Chinese Journal of Digestive Surgery 2023;22(4):519-525
Objective:To investigate the efficacy of different laparoscopic surgeries for gastrointestinal stromal tumors (GIST) of gastric cardia and fundus.Methods:The retrospective cohort study was conducted. The clinicopathological data of 251 patients with GIST of gastric cardia and fundus who underwent laparoscopic radical resection in 14 medical centers, including Guangdong Provincial People′s Hospital et al, from December 2007 to December 2021 were collected. There were 123 males and 128 females, aged 58(24,87)years. Observation indicators: (1) treatment; (2) clinicopathological data of patients undergoing different laparoscopic surgeries; (3) subgroup analysis for special laparoscopic techniques. Measurement data with normal distribution were represented as Mean± SD, and comparison between groups was conducted using the t test or ANOVA. Measure-ment data with skewed distribution were represented as M( Q1, Q3), and comparison between groups was conducted using the Mann-Whitney U test or Kruskal-Wallis H test. Count data were described as absolute numbers or percentages. Comparison of ordinal data was conducted using the rank sum test. Results:(1) Treatment. Of the 251 patients,202 cases underwent gastric wedge resection, 26 cases underwent special laparoscopic techniques including 10 cases with serotomy and dissection and 16 cases with transluminal gastrectomy, 23 cases underwent structural gastrectomy including 6 cases with total gastrectomy and 17 cases with proximal partial gastrectomy. There were 24 patients had postoperative complications after surgery. (2) Clinicopathological data of patients undergoing different laparoscopic surgeries. The gender (male, female), age, tumor diameter, operation time, volume of intraoperative blood loss, length of incision, time to postoperative initial whole liquid food intake, time to postoperative initial semi-liquid food intake, duration of postoperative hospital stay, cases with perioperative complications, cases with mitotic count as ≤5/50 high power field, 6?10/50 high power field, >10/50 high power field, cases be classified as very low risk, low risk, medium risk, high risk according to the National Institutes of Health risk classification, cases with tumor located at fundus and gastric cardia were 93, 109, (59±11)years, 3.50(0.40,10.00)cm, 88.00(25.00,290.00)minutes,20.00(25.00,290.00)mL, 4.00(2.00,12.00)cm, 3.00(1.00,9.00)days, 4.00(1.00,16.00)days, 5.00(1.00,18.00)days, 14, 164, 31, 7, 47, 83, 50, 22, 30, 172 in patients undergoing gastric wedge resection, respectively. The above indicators were 19, 7, (49±14)years, 2.55(0.20,5.00)cm, 101.00(59.00,330.00)minutes, 27.50(2.00,300.00)mL, 4.50(0,6.00)cm, 2.50(1.00,10.00)days, 4.00(1.00,16.00)days, 6.00(1.00,18.00)days, 3, 20, 5, 1, 15, 5, 2, 4, 24, 2 in patients undergoing special laparos-copic techniques, and 11, 12, (52±10)years, 5.00(0.80,10.00)cm, 187.00(80.00,325.00)minutes, 50.00(10.00,300.00)mL, 6.00(4.00,12.00)cm, 4.00(2.00,8.00)days, 6.00(3.00,14.00)days, 8.00(2.00,18.00)days, 7, 11, 5, 7, 2, 6, 6, 9, 13, 10 in patients undergoing structural gastrectomy. There were significant differences in the above indicators among the three groups of patients ( χ2=6.75, F=10.19, H=17.71, 37.50, 35.54, 24.68, 16.09,20.20, 13.76, χ2=13.32, Z=28.98, 32.17, χ2=82.14, P<0.05). (3) Subgroup analysis for special laparoscopic techniques. The time to postoperative initial whole liquid food intake, time to postoperative initial semi-liquid food intake, classification of tumor location (endophytic type, exophytic type, parietal type) were 4.50(1.00,10.00)days, 8.00(3.00,12.00)days, 0, 8, 2 in patients undergoing serotomy and dissection, versus 2.00(1.00,4.00)days, 3.00(1.00,6.00)days, 16, 0, 0 in patients undergoing transluminal gastrectomy. There were significant differences in time to postoperative initial whole liquid food intake, time to postoperative initial semi-liquid food intake between them ( Z=-2.65, -3.16, P<0.05); and there was a significant difference in classification of tumor location between them ( P<0.05). Conclusions:Gastric wedge resection is the most commonly used laparoscopic technique for GIST of gastric cardia and fundus. The application of special laparoscopic techniques is focused on the GIST of cardia to preserve the function of the cardia.
5.Clinical Effect of Chaishao Longmu Decoction on Insomnia in Patients with Liver Depression and Spleen Deficiency Based on Infrared Thermal Images
Langhua ZHENG ; Lichang LIANG ; Xuecheng HUANG ; Jianwen QIU ; Zhiwei ZHANG ; Wei BIN ; Yanzhao LIN
Chinese Journal of Experimental Traditional Medical Formulae 2023;29(23):106-113
ObjectiveTo observe the therapeutic effect of Chaishao Longmu decoction on insomnia in the patients with the syndrome of live depression and spleen deficiency and explore the correlation between infrared thermal imaging and insomnia with liver depression and spleen deficiency. MethodA total of 72 insomnia patients treated in the outpatient department of Shenzhen Hospital of Guangzhou University of Chinese Medicine (Futian) from May to December in 2022 were selected and randomized into a treatment group and a control group, with 36 patients in each group. The patients in the treatment group were treated with Chaishao Longmu decoction and those in the control group with eszopiclone for 4 weeks. The clinical efficacy, Pittsburgh sleep quality index (PSQI) score, TCM syndrome score and infrared thermal imaging characteristics [temperature and temperature changes of frontal sleep line, frontal region, anterior trunk, Zhongwan (CV12), conception vessel (CV), left hypochondrium, right hypochondrium, dorsal trunk, and governor vessel (GV)] of two groups were determined before and after treatment. ResultAfter treatment, the clinical response rate in treatment group was 91.67% (33/36), which was higher than that (66.67%, 24/36) in the control group (Z=-2.617, P<0.01). The treatment in both groups decreased the PSQI score and TCM syndrome score (P<0.01), and the decreases were more significant in treatment group than in the control group (P<0.01). After treatment, the total response rate of sleep line improvement in the treatment group was 86.11% (31/36), which was higher than that (66.67%, 24/36) in the control group (Z=-2.591, P<0.05). The frontal temperature of the two groups decreased (P<0.01) after treatment. Compared with those before treatment, the temperatures of anterior trunk, CV12, CV, left hypochondrium, right hypochondrium, dorsal trunk, and GV rose after treatment (P<0.01). After treatment, the treatment group had lower frontal temperature and higher temperatures of anterior trunk, CV12, CV, left hypochondrium, right hypochondrium, dorsal trunk, and GV than the control group (P<0.01). The treatment in the treatment group reduced the ∆T values of GV (P<0.01) and increased that of the CV12, CV, left hypochondrium, and right hypochondrium (P<0.05,P<0.01). The treatment in the control group increased the ∆T value of CV12 (P<0.01). After treatment, the treatment group had lower ∆T values of GV (P<0.01) and higher ∆T value of CV12, CV, left hypochondrium, and right hypochondrium (P<0.05, P<0.01) than the control group. Compared with that before treatment, the temperature difference between GV and CV in the two groups increased after treatment (P<0.01). According to the infrared thermal image characteristics, the normal temperature difference between GV and CV was within the range of 0.5-1. The median value after treatment in the treatment group was 0.69 (0.52, 0.88), which was within the normal range, indicating that the treatment group outperformed the control group. ConclusionChaishao Longmu decoction can alleviate short-term insomnia by soothing liver, invigorating spleen, harmonizing the middle energizer, and regulating GV and CV. With definite clinical effect, this decoction deserves promotion. Furthermore, the frontal temperature, sleep line, CV12, CV ∆T, and temperature difference between GV and CV revealed by the infrared thermal images could be used for the diagnosis and of insomnia with liver depression and spleen deficiency.
6.Screening and influencing factors of spine problems of children and adolescents aged 6-18 years
Dan YU ; Le ZHAO ; Zhiwei QIU ; Liewei ZHOU ; Tao GONG ; Xuhu ZAN
Journal of Public Health and Preventive Medicine 2023;34(4):135-138
Objective To analyze the screening results of spinal problems in children and adolescents aged 6-18 years and the influencing factors of scoliosis to provide reference for the prevention of spinal problems in children and adolescents. Methods Stratified cluster random sampling was used to screen the prevalence of scoliosis among kindergarten to senior high school students in Shiyan city, and a questionnaire survey was conducted among subjects or parents. Multivariate logistic regression was used to analyze the risk factors affecting the occurrence of scoliosis. Results A total of 1 674 children and adolescents were investigated, and 113 cases of scoliosis were detected, with a detection rate of 6.75%. The probability of scoliosis was 1.92% (13/678), 5.35% (28/523) and 17.76% (72/473) in elementary school, junior high school and senior high school students, respectively. The detection rate of scoliosis gradually increased with the increase of education level (χ2 for trend = 5.272, P < 0.05). In the scoliosis group, the proportions of females (65.49%), malnutrition (25.66%), sitting postural irregularity (52.21%), daily sitting learning time > 12 h (63.72%), daily electronic product use time > 2 h (67.26%), high physical activity > 1 time/d (42.48%) in the past 7 d, and daily outdoor activity time ≤ 2 h (62.83%) were higher than those in the group without scoliosis (P < 0.05).Multivariate logistic regression analysis showed that female students (OR=1.840, 95% CI:1.385-2.716), malnutrition (OR=2.175, 95% CI: 1.542-3.941), improper sitting posture (OR=2.228, 95% CI: 1.592-4.182), daily sitting study time>12 hours (OR=3.258 , 95% CI: 2.562-11.247), daily electronic product use time>2 hours (OR=2.619, 95% CI: 1.935-5.508) , Heavy physical activity in the past 7 days (OR=1.724, 95% CI: 1.347-2.966) , Daily outdoor activity ≤2 h(OR=1.830,95% CI: 1.463-3.103)is a risk factor for scoliosis in children and adolescents (P<0.05). Conclusions The occurrence of scoliosis in children and adolescents is related to gender, nutritional status, and learning habits, and it is necessary to strengthen the screening of high-risk groups in order to reduce the occurrence of scoliosis.
7.MSCs-derived apoptotic extracellular vesicles promote muscle regeneration by inducing Pannexin 1 channel-dependent creatine release by myoblasts.
Qingyuan YE ; Xinyu QIU ; Jinjin WANG ; Boya XU ; Yuting SU ; Chenxi ZHENG ; Linyuan GUI ; Lu YU ; Huijuan KUANG ; Huan LIU ; Xiaoning HE ; Zhiwei MA ; Qintao WANG ; Yan JIN
International Journal of Oral Science 2023;15(1):7-7
Severe muscle injury is hard to heal and always results in a poor prognosis. Recent studies found that extracellular vesicle-based therapy has promising prospects for regeneration medicine, however, whether extracellular vesicles have therapeutic effects on severe muscle injury is still unknown. Herein, we extracted apoptotic extracellular vesicles derived from mesenchymal stem cells (MSCs-ApoEVs) to treat cardiotoxin induced tibialis anterior (TA) injury and found that MSCs-ApoEVs promoted muscles regeneration and increased the proportion of multinucleated cells. Besides that, we also found that apoptosis was synchronized during myoblasts fusion and MSCs-ApoEVs promoted the apoptosis ratio as well as the fusion index of myoblasts. Furthermore, we revealed that MSCs-ApoEVs increased the relative level of creatine during myoblasts fusion, which was released via activated Pannexin 1 channel. Moreover, we also found that activated Pannexin 1 channel was highly expressed on the membrane of myoblasts-derived ApoEVs (Myo-ApoEVs) instead of apoptotic myoblasts, and creatine was the pivotal metabolite involved in myoblasts fusion. Collectively, our findings firstly revealed that MSCs-ApoEVs can promote muscle regeneration and elucidated that the new function of ApoEVs as passing inter-cell messages through releasing metabolites from activated Pannexin 1 channel, which will provide new evidence for extracellular vesicles-based therapy as well as improving the understanding of new functions of extracellular vesicles.
Creatine/metabolism*
;
Extracellular Vesicles
;
Muscle, Skeletal/metabolism*
;
Myoblasts/metabolism*
;
Regeneration
;
Connexins/metabolism*
8.Research progress on AMPK activation in nonalcoholic fatty liver disease
Xinyu YE ; Tongxi LI ; Zhiwei HUANG ; Wenguang FU ; Qiu LI
Chinese Journal of Hepatology 2023;31(11):1220-1223
Nonalcoholic fatty liver disease (NAFLD) occurrence and progression are associated with lipid accumulation, insulin resistance, inflammation, liver damage, fibrosis, and other factors. AMP-dependent protein kinase (AMPK) is a key molecule that regulates bioenergy metabolism and participates in multiple biological processes, including lipid metabolism, autophagy, inflammation, and cell apoptosis. Promoting AMPK activation can reduce hepatic lipid accumulation and insulin resistance, alleviate the development of NAFLD, reduce liver inflammation and fibrosis, and inhibit the progression of NAFLD to nonalcoholic steatohepatitis.
9.The clinical study of pelvic sacral acupuncture combined with myofascial manipulation in the treatment of postpartum pelvic pain
Jiao CHENG ; Congshan CHEN ; Yuanyang ZHANG ; Zhiwei QIU ; Yan WANG ; Xiaoyong ZHANG
International Journal of Traditional Chinese Medicine 2022;44(11):1243-1247
Objective:To evaluate the efficacy of pelvic sacral tendon regulating acupuncture combined with myofascial manipulation in the treatment of postpartum pelvic pain.Methods:A total of 86 patients with postpartum pelvic pain in our hospital from December 2019 to October 2021 who met the inclusion criteria were divided into control group ( n=44) and observation group ( n=42) by random number table method. The control group was treated with myofascial manipulation, while the observation group was treated with pelvic sacral tendon acupuncture on the basis of the control group. Both groups were treated for 4 weeks. The VAS was used to evaluate the degree of pain, Oswestry Disability Index (ODI) to evaluate the degree of lumbar dysfunction, pelvic floor dysfunction Questionnaire 7 (PFIQ-7) to evaluate the pelvic floor function, and pelvic floor dysfunction Questionnaire (PFDI20) to evaluate the quality of life. Serum levels of insulin-like growth factor-1 (IGF-1) and TGF-β1 were detected by ELISA to evaluate the clinical efficacy. Results:The total effective rate was 95.24% (40/42) in the observation group and 79.55% (35/44) in the control group, and the difference was statistically significant ( χ2=4.74, P=0.030). After treatment, the ODI, VAS, PFIQ-7 and PFIQ-20 scores of the observation group were significantly lower than those in the control group ( t=13.79, 14.05, 10.61, 10.10, all Ps<0.001). The scores of low back pain, limb pain and gait claudication were significantly lower than those in the control group ( t=11.73, 12.16 and 10.89, respectively, all Ps<0.001). The serum levels of TGF-β1 [(163.81±28.05) μg/L vs. (124.56±19.23) μg/L, t=5.76] and IGF-1 [(64.52±9.14) μg/L vs. (49.86±7.62) μg/L, t=5.06] were significantly higher than those in the control group ( P<0.01). Conclusion:The pelvic sacral tendon acupuncture combined with myofascial manipulation can improve the pelvic function of patients with postpartum pelvic pain, relieve the degree of pelvic pain, and improve the curative effect.
10.Clinical features and genetic analysis of a Chinese pedigree affected with X-linked adrenoleukodystrophy.
Lingen KONG ; Jinhua QIU ; Caiming LI ; Qianwu ZHOU ; Zhiwei QIU ; Shunzhi ZHUANG ; Jinxiu ZOU ; Yanxia ZHENG
Chinese Journal of Medical Genetics 2022;39(1):60-63
OBJECTIVE:
To analyze the clinical features and variants of ABCD1 gene in a Chinese pedigree affected with X-linked adrenoleukodystrophy.
METHODS:
Clinical data of the proband were collected and analyzed. Potential variant of the ABCD1 gene were analyzed by PCR and Sanger sequencing of the proband, his parents and 100 unrelated healthy individuals.
RESULTS:
The prominent features of the proband included cerebellar and brainstem lesions, along with increased serum level of very-long chain fatty acids. He was found to harbor a hemizygous c.1509delG (p.L504Sfs*54) variant of the ABCD1 gene, for which his mother was heterozygous. The same variant was not detected in his father and 100 healthy controls.
CONCLUSION
X-linked adrenoleukodystrophy has a variety of clinical manifestations. Discovery of the c.1509delG (p.L504Sfs*54), as a novel pathogenic variant of the ABCD1 gene, has enabled diagnosis and genetic counseling for this pedigree.
Adrenoleukodystrophy/genetics*
;
Asians/genetics*
;
China
;
Female
;
Genetic Testing
;
Humans
;
Male
;
Mutation
;
Pedigree


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