1.Integrated Transcriptomic Landscape and Deep Learning Based Survival Prediction in Uterine Sarcomas
Yaolin SONG ; Guangqi LI ; Zhenqi ZHANG ; Yinbo LIU ; Huiqing JIA ; Chao ZHANG ; Jigang WANG ; Yanjiao HU ; Fengyun HAO ; Xianglan LIU ; Yunxia XIE ; Ding MA ; Ganghua LI ; Zaixian TAI ; Xiaoming XING
Cancer Research and Treatment 2025;57(1):250-266
Purpose:
The genomic characteristics of uterine sarcomas have not been fully elucidated. This study aimed to explore the genomic landscape of the uterine sarcomas (USs).
Materials and Methods:
Comprehensive genomic analysis through RNA-sequencing was conducted. Gene fusion, differentially expressed genes (DEGs), signaling pathway enrichment, immune cell infiltration, and prognosis were analyzed. A deep learning model was constructed to predict the survival of US patients.
Results:
A total of 71 US samples were examined, including 47 endometrial stromal sarcomas (ESS), 18 uterine leiomyosarcomas (uLMS), three adenosarcomas, two carcinosarcomas, and one uterine tumor resembling an ovarian sex-cord tumor. ESS (including high-grade ESS [HGESS] and low-grade ESS [LGESS]) and uLMS showed distinct gene fusion signatures; a novel gene fusion site, MRPS18A–PDC-AS1 could be a potential diagnostic marker for the pathology differential diagnosis of uLMS and ESS; 797 and 477 uterine sarcoma DEGs (uDEGs) were identified in the ESS vs. uLMS and HGESS vs. LGESS groups, respectively. The uDEGs were enriched in multiple pathways. Fifteen genes including LAMB4 were confirmed with prognostic value in USs; immune infiltration analysis revealed the prognositic value of myeloid dendritic cells, plasmacytoid dendritic cells, natural killer cells, macrophage M1, monocytes and hematopoietic stem cells in USs; the deep learning model named Max-Mean Non-Local multi-instance learning (MMN-MIL) showed satisfactory performance in predicting the survival of US patients, with the area under the receiver operating curve curve reached 0.909 and accuracy achieved 0.804.
Conclusion
USs harbored distinct gene fusion characteristics and gene expression features between HGESS, LGESS, and uLMS. The MMN-MIL model could effectively predict the survival of US patients.
2.Current status of eating behaviors and its predictive role in overweight and obese of adolescents
Chinese Journal of School Health 2025;46(1):53-57
Objective:
To explore the current status and influencing factors of eating behaviors in adolescents, so as to provide a theoretical foundation for health promotion education among adolescents.
Methods:
Based on the database from Survey of Chinese Family Health Index (2021), by a random number table method, 1 065 teenagers were selected from the provincial capitals of 22 provinces and 5 autonomous regions in China, as well as 4 municipalities directly under the central government. A general characteristic questionnaire, Patient Health Questionnaire-9 (PHQ-9), Short Form of the Family Health Scale (FHS-SF), 10-item Short Version of the Big Five Personality(BFP-10), Content-based Media Exposure Scale (CM-E) and Sakata Eating Behavior Scale Short Form(EBS-SF) were used to collect information. Multivariate stepwise linear regression analysis was employed to identify and analyze related factors of eating behaviors among adolescents. Receiver operating characteristic was used to validate the predictive ability of the EBS-SF score for overweight and obesity among adolescents.
Results:
The average scores of BFI-10,C-ME, FHS-SF, PHQ-9 and EBS-SF were (33.08±4.64)(19.20±4.55)(38.48±6.65)(6.09±5.63)(16.75±4.36), respectively. Multivariate linear regression showed that family type (other types), agreeableness, conscientiousness, family health and depression were the main related factors of EBS-SF scores among adolescents( B =2.61,-0.42,0.20,-0.11,0.23, P <0.05).The analysis of receiver operating characteristic curve showed that the EBS-SF scores had a good ability in predicting obesity among male adolescents ( AUC= 0.73, P <0.01).
Conclusions
Family type, big five personality, family health,depression are the related factors of eating behaviors among adolescents. EBS-SF scores are predictive of obesity in adolescents, which would provide a new perspective for promoting healthy eating habits among adolescents.
3.Integrated Transcriptomic Landscape and Deep Learning Based Survival Prediction in Uterine Sarcomas
Yaolin SONG ; Guangqi LI ; Zhenqi ZHANG ; Yinbo LIU ; Huiqing JIA ; Chao ZHANG ; Jigang WANG ; Yanjiao HU ; Fengyun HAO ; Xianglan LIU ; Yunxia XIE ; Ding MA ; Ganghua LI ; Zaixian TAI ; Xiaoming XING
Cancer Research and Treatment 2025;57(1):250-266
Purpose:
The genomic characteristics of uterine sarcomas have not been fully elucidated. This study aimed to explore the genomic landscape of the uterine sarcomas (USs).
Materials and Methods:
Comprehensive genomic analysis through RNA-sequencing was conducted. Gene fusion, differentially expressed genes (DEGs), signaling pathway enrichment, immune cell infiltration, and prognosis were analyzed. A deep learning model was constructed to predict the survival of US patients.
Results:
A total of 71 US samples were examined, including 47 endometrial stromal sarcomas (ESS), 18 uterine leiomyosarcomas (uLMS), three adenosarcomas, two carcinosarcomas, and one uterine tumor resembling an ovarian sex-cord tumor. ESS (including high-grade ESS [HGESS] and low-grade ESS [LGESS]) and uLMS showed distinct gene fusion signatures; a novel gene fusion site, MRPS18A–PDC-AS1 could be a potential diagnostic marker for the pathology differential diagnosis of uLMS and ESS; 797 and 477 uterine sarcoma DEGs (uDEGs) were identified in the ESS vs. uLMS and HGESS vs. LGESS groups, respectively. The uDEGs were enriched in multiple pathways. Fifteen genes including LAMB4 were confirmed with prognostic value in USs; immune infiltration analysis revealed the prognositic value of myeloid dendritic cells, plasmacytoid dendritic cells, natural killer cells, macrophage M1, monocytes and hematopoietic stem cells in USs; the deep learning model named Max-Mean Non-Local multi-instance learning (MMN-MIL) showed satisfactory performance in predicting the survival of US patients, with the area under the receiver operating curve curve reached 0.909 and accuracy achieved 0.804.
Conclusion
USs harbored distinct gene fusion characteristics and gene expression features between HGESS, LGESS, and uLMS. The MMN-MIL model could effectively predict the survival of US patients.
4.Integrated Transcriptomic Landscape and Deep Learning Based Survival Prediction in Uterine Sarcomas
Yaolin SONG ; Guangqi LI ; Zhenqi ZHANG ; Yinbo LIU ; Huiqing JIA ; Chao ZHANG ; Jigang WANG ; Yanjiao HU ; Fengyun HAO ; Xianglan LIU ; Yunxia XIE ; Ding MA ; Ganghua LI ; Zaixian TAI ; Xiaoming XING
Cancer Research and Treatment 2025;57(1):250-266
Purpose:
The genomic characteristics of uterine sarcomas have not been fully elucidated. This study aimed to explore the genomic landscape of the uterine sarcomas (USs).
Materials and Methods:
Comprehensive genomic analysis through RNA-sequencing was conducted. Gene fusion, differentially expressed genes (DEGs), signaling pathway enrichment, immune cell infiltration, and prognosis were analyzed. A deep learning model was constructed to predict the survival of US patients.
Results:
A total of 71 US samples were examined, including 47 endometrial stromal sarcomas (ESS), 18 uterine leiomyosarcomas (uLMS), three adenosarcomas, two carcinosarcomas, and one uterine tumor resembling an ovarian sex-cord tumor. ESS (including high-grade ESS [HGESS] and low-grade ESS [LGESS]) and uLMS showed distinct gene fusion signatures; a novel gene fusion site, MRPS18A–PDC-AS1 could be a potential diagnostic marker for the pathology differential diagnosis of uLMS and ESS; 797 and 477 uterine sarcoma DEGs (uDEGs) were identified in the ESS vs. uLMS and HGESS vs. LGESS groups, respectively. The uDEGs were enriched in multiple pathways. Fifteen genes including LAMB4 were confirmed with prognostic value in USs; immune infiltration analysis revealed the prognositic value of myeloid dendritic cells, plasmacytoid dendritic cells, natural killer cells, macrophage M1, monocytes and hematopoietic stem cells in USs; the deep learning model named Max-Mean Non-Local multi-instance learning (MMN-MIL) showed satisfactory performance in predicting the survival of US patients, with the area under the receiver operating curve curve reached 0.909 and accuracy achieved 0.804.
Conclusion
USs harbored distinct gene fusion characteristics and gene expression features between HGESS, LGESS, and uLMS. The MMN-MIL model could effectively predict the survival of US patients.
5.Mechanism of warmed malate ringer's solution in fluid resuscitation in improving the lethal triad of severe trauma
Yinyu WU ; Han SHE ; Yunxia DU ; Yuxi ZHANG ; Xiaowei ZHOU ; Qinghui LI ; Tao LI ; Yi HU ; Qingxiang MAO ; Yaling WANG
Journal of Army Medical University 2025;47(3):216-225
Objective To explore the role and mechanism of warm malate ringer's solution(MR)in resuscitation of the lethal triad caused by severe trauma.Methods A rat model of severe trauma was established in SPF-grade SD rats(half male and half female,weighing 200~220 g)using combined multiple injuries and hemorrhagic shock,and the rats were randomly divided into 8 groups(n=8):Sham group,only arterial and venous catheterization;Trauma(Tra)groups with different time points(10,30,60,90,120,180 min)and a Trauma group that were observed without any treatment for 180 min after model establishment.The changes of activated clotting time(ACT),reaction time(R),maximum amplitude(MA),and rate of blood clot formation(Angle)at different time points were detected by using thromboelastography,and tail bleeding,core body temperature and arterial blood gas parameters,were also observed and detected.The plasma von Willebrand Factor(vWF)level,mitochondrial respiratory control ratio in pulmonary venous endothelium,and expression levels of vascular endothelial cadherin(VE-Cadherin),peroxisome proliferator activating receptor gamma coactivator 1α(PGC1α),dynamin-related protein 1(Drp1),p-Drp1,and mitofusin 2(Mfn2)were detected to evaluate the vascular endothelial injury and mitochondrial dysfunction.Another group of SD rats were randomly divided into severe trauma group(no treatment for 180 min after injury),and MR solution at room temperature and at 37 ℃ groups.MR solution at room temperature or at 37 ℃ was given to the rats using a medical blood transfusion apparatus at 60 min post-trauma.Above indicators were observed and detected to investigate the resuscitation effect of the MR solution.Results Compared with the Sham group,the severely traumatic rats at 180 min after injury had significantly prolonged ACT and R values(P<0.05),shortened MA and decreased Angle values(P<0.05),extended tail bleeding time(P<0.05),lower partial pressure of carbon dioxide(PCO2)and HCO3-and base excess(BE)levels(P<0.05),and continuously increasing K+(P<0.05)and decreasing Na+(P<0.05)and Ca2+levels(P<0.05).Additionally,plasma vWF level(P<0.05)and protein levels of VE-cadherin,PGC1α and Mfn2 in pulmonary vein endothelium were significantly reduced(P<0.05),the expression of p-Drp1 was enhanced and the mitochondrial respiration control rate was declined in the rats at 180 min after injury(P<0.05).MR solution resuscitation shortened tail bleeding time(P<0.05),increased core body temperature(P<0.05),elevated plasma vWF level(P<0.05),increased protein levels of VE-cadherin,PGC1α and Mfn2(P<0.05),and decreased that of p-Drp1 protein expression(P<0.05)when compared with the rats at 180 min after severe traumatic injury.The above effects were more significant in the rats infused with the solution at 37 ℃ than those at room temperature.Conclusion Warm MR solution significantly improves the lethal triad in rats after severe trauma,which may be associated with its improving mitochondrial function and attenuating vascular endothelial damage.
6.Clinical characteristics of neonatal necrotizing enterocolitis and analysis of risk factors for early-onset children
Jing WANG ; Mingqi SHEN ; Rongxiu ZHENG ; Yue XIN ; Yunxia MA ; Ying ZHANG ; Dejing WU ; Dan LIU
International Journal of Pediatrics 2025;52(9):629-633
Objective:To explore the clinical characteristics of neonatal necrotizing enterocolitis(NEC)and analyze the risk factors for early-onset NEC.Methods:A total of 220 children with NEC admitted to the Department of Pediatrics,Tianjin Medical University General Hospital from January 1st,2018 to February 29th,2024 were retrospectively selected as the research objects. According to the time of onset,the early-onset group( n=120)and the late-onset group( n=100)were established,and the clinical characteristics of the two groups were compared. Another 150 cases of normal healthy newborns born in this hospital in the same period were selected as the control group,and the clinical data of the control group were collected. The clinical characteristics of the early-onset group and the control group were compared,and the risk factors of early-onset NEC were analyzed by multivariate Logistic regression. Results:Compared with the late-onset group,the early-onset group had fever[50.0%(60/120)vs. 40%(40/100), χ2=7.333, P=0.007],apnea[39.17%(47/120)vs. 28%(28/100), χ2=7.568, P=0.006],no rise in body temperature[56.67%(68/120)vs. 39%(39/100), χ2=6.815, P=0.009],abdominal distension[25%(30/120)vs. 40%(40/100), χ2=13.200, P<0.001],vomiting[30.83%(37/120)vs. 45%(45/100), χ2=12.797, P<0.001]was significantly different(all P<0.05);Multivariate Logistic regression analysis:weight<1 500 g( OR=5.871,95% CI:3.153~9.673, P<0.001),gestational age<30 weeks( OR=4.256,95% CI:2.641~7.896, P=0.007),hemodynamically significant patent ductus arteriosus(hs-PDA)( OR=3.113,95% CI:1.865~5.133, P=0.033),severe anemia( OR=3.057,95% CI:2.165~4.802, P=0.001),feeding intolerance( OR=4.215,95% CI:1.579~10.802, P=0.005),amniotic fluid pollution( OR=2.452,95% CI:1.579~3.111, P<0.001)were the independent risk factors for early-onset NEC(all P<0.05). Conclusion:Weight<1 500 g,gestational age<30 weeks,hs-PDA,severe anemia,feeding intolerance,and amniotic fluid contamination are independent risk factors for early-onset NEC. In clinical practice,more attention should be paid to these factors for disease prevention,early identification,and timely intervention in newborns to reduce the occurrence of NEC.
7.Clinical observation of regional citrate anticoagulation in pediatric plasma exchange
Wei DANG ; Fan ZHANG ; Yunxia LI ; Jie CHEN ; Xia LIN ; Sufang ZHANG ; Weifeng LU
Chinese Journal of Emergency Medicine 2025;34(6):795-802
Objective:To evaluate the efficacy and safety of regional citrate anticoagulation (RCA) during plasma exchange (PE) in pediatric patients.Methods:We conducted a retrospective analysis of 12 critically ill children admitted to the Pediatric Intensive Care Unit (PICU) of Jinan Children's Hospital, who underwent 28 PE sessions with RCA between December 2023 and August 2024. Clinical records were reviewed to assess bleeding events, extracorporeal circuit performance, and changes in arterial blood gas parameters, serum total calcium (Ca tot), and activated clotting time before and after treatment. Results:No patients exhibited signs of increased bleeding. In one case, the procedure was discontinued prematurely due to elevated venous pressure. A significant decrease in ionized calcium (Ca ion) was observed 0.5 hours post-treatment. At the end of PE, pH, HCO 3?, base excess (BE), lactate, PaCO 2, Ca tot, and Na + levels increased, while K + and Ca ion levels decreased, with all changes being statistically significant. Four hours post-treatment, pH, HCO 3?, BE, PaCO 2, and Na + remained elevated, whereas Ca ion, lactate, and K + returned to baseline. By 12–15 hours post-treatment, all parameters—including pH, HCO 3?, BE, PaCO 2, Na +, K +, Ca ion, and lactate—had normalized, showing no significant differences from pre-treatment levels. Conclusions:RCA provides effective extracorporeal anticoagulation during pediatric PE without increasing bleeding risk. However, metabolic complications—primarily metabolic alkalosis—are common. These disturbances typically resolve spontaneously and do not lead to severe adverse events. While no ideal anticoagulant for PE has yet been established, RCA remains a safe and effective option, particularly for pediatric patients at higher risk of bleeding.
8.Construction and application of an evidence-based practice plan for early activity in postoperative patients with cerebral infarction
Yunxia YU ; Chun ZHANG ; Lijuan ZHU
Chinese Journal of Practical Nursing 2024;40(4):289-295
Objective:To explore the construction and application of an evidence-based practice plan for early postoperative activity in postoperative patients with large area cerebral infarction.Methods:Ninety-six postoperative patients with large area cerebral infarction admitted to Wenzhou Central Hospital from July 2021 to April 2023 were selected as the study subjects for a Clinical trial. They were divided into the control group (48 cases) and the observation group (48 cases) by random number table method. The control group received routine postoperative care for neurosurgery, while the observation group received evidence-based systematic early activity training. Both groups were followed up until 1 month after the patient was discharged from the hospital. The time of postoperative hospitalization, hospitalization expenses, vital signs and pain investigation 48 h after surgery, daily living ability before intervention and 7 d, 1 month after discharge, neurological function before intervention and 7 d and 1 month after surgery, and complications during follow-up between the two groups were compared.Results:In the control group, there were 29 males and 19 females, with an average age of 43-67(56.87 ± 1.76) years. In the observation group, there were 31 males and 17 females, with an average age of 43-68 (57.02 ± 1.82) years. The postoperative hospital stay in the observation group was (6.87 ± 0.65) d, in the control group was (9.06 ± 0.72) d, the difference between them was significant ( t=15.64, P<0.05). 48 hours after surgery, the heart rate, breathing rate and mean arterial pressure in the observation group were (71.65 ± 0, 45) times/min, (14.76 ± 0.36) times/min and (76.98 ± 5.65) mmHg(1 mmHg=0.133 kPa), which were different with those in the control group, (82.76 ± 2.65) times/min, (18.76 ± 2.87) times/min and (93.76 ± 5.93) mmHg ( t=28.64, 9.58 and 14.19, all P<0.05). Seven days after discharge, the score of Activities of Daily Living and the National Institutes of Health Neurological Impairment Scale in the observation group were (84.65 ± 2.45) and (23.65 ± 2.65), which were different than the (79.76 ± 1.97) and (28.54 ± 2.73) in the control group ( t=10.26, 8.91, both P<0.05). A month after discharge, the score of Activities of Daily Living and the National Institutes of Health Neurological Impairment Scale in the observation group were (95.45 ± 1.43) and (18.65 ± 1.98), while in the control group were (87.87 ± 1.39) and (21.54 ± 2.76), the difference between them were significant ( t=26.33, 5.90, both P<0.05). The total incidence of complications such as postoperative bleeding, postoperative infection and hypoxemia in the observation group was 20.83% (10/48), which was significant lower than the 68.75% (33/48) in the control group ( χ2=22.28, P<0.05). Conclusions:Evidence-based systematic early activity training could significantly alleviate postoperative pain in patients with large area cerebral infarction after surgery, improve their daily living ability and neurological function, further effectively shorten their hospitalization time, and reduce the occurrence of complications.
9.Application research of PGT in blocking the inheritance of novel mutations in the PKHD1 gene in autoso-mal recessive polycystic kidney disease pedigrees
Ning WANG ; Yan HAO ; Dawei CHEN ; Zhiguo ZHANG ; Dan KUANG ; Qing ZHANG ; Yiqi YING ; Zhaolian WEI ; Ping ZHOU ; Yunxia CAO
The Journal of Practical Medicine 2024;40(7):1006-1010
Objective To investigate the application value of single nucleotide polymorphism(SNP)linkage analysis based on next-generation sequencing(NGS)technology in preimplantation genetic testing(PGT)of families with autosomal recessive polycystic kidney disease(ARPKD).Methods A family with ARPKD was selected,where the female member had a pregnancy ultrasound revealing polycystic kidney in the fetus.Genetic testing showed compound heterozygous mutations of the polycystic kidney/polycystic liver disease 1 gene(PKHD1),c.10444C>T(paternal)and c.4303del(maternal),with the c.4303del mutation being reported for the first time.Targeting the coding region of the PKHD1 gene,335 high-density tightly linked SNP sites were selected in the upstream and downstream 2M regions using multiplex polymerase chain reaction(PCR)and NGS.The couple′s SNP risk haplotypes carrying gene mutations were constructed.After in vitro fertilization,blastocyst culture was performed.Trophoblastic cells obtained from the biopsy were subjected to whole-genome amplification,and NGS was used for linkage analysis and low-depth chromosomal aneuploidy screening of the embryos.Sanger sequencing was used to verify the results of embryo linkage analysis.Results Among the 6 biopsied embryos,4 were mutation-free and euploid,1 exhibited heterozygous for the mutation and mosaic while another unstable sequencing data,making it impossible to judge.One of the mutation-free and developmentally healthy euploid embryos was implanted into the maternal uterus,resulting in the full-term delivery of a healthy baby.Conclusion Application of NGS-based SNP linkage analysis in PGT can effectively blocking the vertical transmission of ARPKD within families,while avoiding abortion issues caused by aneuploid embryos.This study is also the first PGT report target-ing the PKHD1 gene c.4303del mutation.
10.Strength of association between follicular fluid melatonin levels and pregnancy rates in single-cycle in vitro fertilization-embryo transfer women
Shanshan LIU ; Juan WU ; Change CHEN ; Yunxia CAO ; Zhiguo ZHANG
Chinese Journal of Tissue Engineering Research 2024;28(31):4975-4979
BACKGROUND:In vitro fertilization-embryo transfer is commonly used to solve infertility,but its success rate is not high,the more common reasons are poor endometrial receptivity,poor egg quality,etc.The follicular fluid melatonin can inhibit the aging of the ovary,to a certain extent,can promote the development of embryos,improve the probability of conception,but whether there is a correlation between the two is not known. OBJECTIVE:To explore the correlation between follicular fluid melatonin level and pregnancy rate of single-cycle in vitro fertilization-embryo transfer women. METHODS:A total of 112 female patients who received in vitro fertilization-embryo transfer treatment in the First Affiliated Hospital of Anhui Medical University from December 2020 to April 2021 were selected as the study subjects.They were divided into quartile array(Q1-Q5)according to the follicular fluid melatonin level from low to high.Among them,the melatonin level of group Q1 was<6.99 ng/L(n=18),that of group Q2 was 7.00-9.99 ng/L(n=26),that of group Q3 was 10.00-11.99 ng/L(n=27),and that of group Q4 was 12.00-13.99 ng/L(n=18);and melatonin levels in group Q5 were 14.00-19.99 ng/L(n=23).Clinical data characteristics of the five groups were compared.Multi-factor Logistic regression was used to analyze the correlation between follicular fluid melatonin level and pregnancy rate of women with single-cycle in vitro fertilization-embryo transfer and embryo transfer.A restricted cubic spline Logistic regression model was established to analyze the dose-response relationship,and the model was evaluated by clinical decision curve. RESULTS AND CONCLUSION:(1)Compared with the study population with the lowest melatonin quintile(Q1),with the increase of melatonin level(Q2-Q5),the levels of egg harvest and pregnancy success were gradually increased,and the body mass index was gradually decreased,and the differences were significant(P<0.05).(2)Multivariate Logistic regression analysis showed that after adjusting for confounding factors such as global mass index,number of eggs retrieved,luteinizing hormone,estradiol,progesterone and other confounding factors,follicular fluid melatonin level was still independently correlated with pregnancy rate of single-cycle in vitro fertilization-embryo transfer women(OR=1.538,95%CI:1.032-1.837,P<0.05),and there was significant difference in trend test of follicular fluid melatonin level from low to high quintile array(Ptrend<0.05).(3)The sensitivity test analysis showed that E value was 2.117.Subgroup analysis showed that the study population with higher levels of luteinizing hormone in follicular fluid had a more significant association between follicular fluid melatonin and pregnancy rate in single-cycle in vitro fertilization-embryo transfer women(P interaction=0.008).(4)The results of restricted cubic spline model analysis showed that there was a nonlinear dose-response relationship between follicular fluid melatonin level and pregnancy rate of single-cycle in vitro fertilization-embryo transfer women(P<0.05),and there was an overall positive correlation between follicular fluid melatonin level and pregnancy rate of single-cycle in vitro fertilization-embryo transfer women.(5)The results of clinical decision curve analysis showed that the follicular fluid melatonin level had important clinical value in predicting the pregnancy rate of single-cycle in vitro fertilization-embryo transfer women.(6)Follicular fluid melatonin level is closely related to the pregnancy rate of single-cycle in vitro fertilization-embryo transfer women,and with the decrease of follicular fluid melatonin level,the pregnancy rate of single-cycle in vitro fertilization-embryo transfer women also decreases.


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