Objective:To study the gene regulatory network during the osteogenic differentiation process of dental follicle stem cells (DFSCs)with bioinformatic analysis.Methods:The differentially expressed genes during the osteogenic diffrentiation process of DF-SCs were selected from the GEO Datasets.Then the relationship among the genes were analysed using DAVID and GeneMANIA data-base.Results:Numerous differentially expressed genes during the osteogenic differentiation process of DFSCs were obtained,289 genes with significance of different expression(P <0.05)were enriched into “cell differentiation”,“cell proliferation”,“skeletal sys-tem development”and “calcium signaling pathway”subgroups.ALPL,CTSK,TUFT1 ,TGFBR2,FHL2,ROR2,STC1 ,POSTN, ZBTB1 6,FRZB,PRELP and IGFBP5 were enriched into the “skeletal system development”subgroup.The 1 2 genes exhibited interac-tions,including co-expression,co-localization,genetic interaction and signal pathways.Conclusion:There is a complex molecular regulatory network among the differentially expressed genes during the osteogenic differentiation process of DFSCs.It is necessary to an-alyse the osteogenic differentiation process of DFSCs as a whole from WNT,TGFbeta,MAPK siganal pathyways,Homeobox transcrip-tion factors and osteogenic differentiation marker genes.

Objective To investigate the immune response of the fusions of ovarian carcinoma cells to dendritic cell(DC) transfected with interleukin(IL)-12 gene in vitro. Methods Recombinant human granulocyte macrophage colony stimulating factor and recombinant human tumor necrosis factor ? were used to generate DC from cord blood in vitro. IL-12 fusion gene was cloned into a eukaryotic vector-pcDNA3.1(+). DC were transfected with IL-12-pcDNA3.1(+) using lipofectamine transfection method and electric transfection method respectively. Then polyethylene glycol mediated the fusion of transfected DC and ovarian carcinoma cells, and 3-(4,5-dimethylthiazol-2-yl) 2,5-diphenyl tetrazolium bromide (MTT) test was carried out to observe the immune response. Results DC were generated in vitro from cord blood and expressed high levels of costimulatory(50%) and MHC Ⅱ molecules(99%). RT-PCR showed that IL-12-pcDNA3.1(+) had been successfully transfected into DC. RT-PCR and immunofluorescence analysis showed that DC were fused with ovarian carcinoma cells successfully. Then the fusion cells of ovarian carcinoma cells to transfected DC, and the fusions of ovarian carcinoma cells to DC were cocultured with peripheral blood mononuclear cell respectively. MTT test showed that both fusions could induce cytolytic T cell activity and lysis of ovarian carcinoma cells,and the former effect was stronger. Conclusion The cytolytic T cell activity induced by the fusions of ovarian carcinoma cells to transfected DC is enhanced.

Objective Detecting plasma level of circular RNA(circRNA)hsa_circ_0009024 in pulmonary tuberculosis(TB)patients,and evaluating its diagnostic value for TB.Methods From January 2016 to December 2016, a hosptial-based, case-control study was performed, which include 90 untreated active pulmonary tuberculosis patients(TB group),75 healthy people(healthy control)and 84 patient with other diseases(other disease group).Plasma level of circRNA hsa_circ_0009024 was detected by real-time quantitative polymerase chain reaction.Furthermore, the 90 patients with TB were divided into different subgroups according to cavity formation and the lung fields involvement: patients without lung cavity(55 cases)vs those with lung cavity(35 cases),patients with involvement of <2 lung fields(49 cases)vs≥2 lung fields(41 cases).Plasma levels of hsa _circ_0009024 of 41 TB patientswere monitored andcomparedbefore and after 3 months anti-TB therapy.The sensitivity and specificity of plasma hsa_circ_0009024 were analyzed by using the receiver operating characteristic(ROC)curve.The comparison between two groups was performed with Mann-Whitney U test and the comparison among multigroupswas conducted with Kruskal-Wallis H test.Results Plasma levels of hsa_circ_0009024 in TB patients[1.98(1.42, 2.71)]were significantly higher than healthy controls[1.03(0.78,1.33)]and other disease groups[1. 13(0.77,1.51)](H=76.58,P<0.0001).Plasma levels of hsa_circ_0009024 in cavity pulmonary TB patients were higher than pulmonary TB patients without cavity(U=392.50,P<0.0001).Plasma levels of hsa_circ_0009024 in TB patients with involvement of ≥2 lung fields were higher than <2 lung fields(U=590.50,P=0.0008).As compared to pre -treatment[2.01(1.41, 2.71)], the plasma hsa_circ_0009024 levels decreased significantly in 3 months[1.22(0.85,1.47)](U=292.50,P<0.0001)after anti-TB therapy.The area under the receiver operating characteristics curve(AUC)of plasma hsa_circ_0009024 in discriminating the patients with TB from normal controls, pneumonia patients and lung cancer patients were 0.841 and 0.811, respectively.Conclusion The hsa_circ_0009024 can be used as a potential biomarker in TB diagnosis and monitoring.

Objective:To compare the value of NoSAS score, STOP-BANG questionnaire (SBQ) and Epworth Sleepiness Scale (ESS) in assessing the risk of obstructive sleep apnea hypopnea syndrome (OSAHS) in patients with respiratory disease (RD).Methods:The clinical data, NoSAS, SBQ and ESS scores of 190 patients who underwent overnight polysomnography (PSG) were collected. According to the receiver operating characteristic (ROC) curve, with different apnea-hypopnea index (AHI) as the judgment cutoff, the sensitivity, specificity, positive predictive value, negative predictive value, diagnostic odds ratio (DOR) and accuracy of the three scales were compared.Results:With AHI ≥5 times/h as the cutoff, the area under the ROC curve (AUC) of NoSAS and SBQ were 0.833 and 0.729, respectively, indicating that both have predictive value for mild OSAHS. Among them, NoSAS had a larger DOR value (16.150), indicating that NoSAS had the higher accuracy in assessing the risk of mild OSAHS. When AHI>15 times/h was used as the cutoff, the AUC value of NoSAS was 0.704, indicating that it has predictive value for moderate OSAHS. When AHI>30 times/h was used as the cutoff, the AUC value (0.706) and DOR value (6.527) of SBQ were high, indicating that it has predictive value and good accuracy for severe OSAHS. The SBQ is more sensitive than NoSAS and ESS when evaluating patients at high risk for OSAHS ( SBQ≥3). Conclusions:When evaluating the risk of mild and moderate OSAHS in RD patients, NoSAS is better than SBQ and ESS, and when evaluating severe OSAHS, SBQ is better than NoSAS and ESS. In clinical work, appropriate predictive tools should be selected according to the actual situation to assess the risk of OSAHS, so as to formulate and implement early intervention plans based on the assessment results.

Objective: To investigate the pathologic features of gonadal tissues of disorders of sexual development (DSD) in children. Methods: Fifty-three cases of gonadal developmental disorders were collected from July 2015 to August 2017 at Guangzhou Women and Children′s Medical Center. Clinical manifestations, karyotypes, sex hormone levels, ultrasound imaging, histology and immunophenotype of gonadal tissues were analyzed. Results: The age of patients ranged from 7 months to 17 years with an average of (50.7 ± 47.1) months. Social genders of the patients included 32 males and 21 females. Forty-eight patients had abnormal sex hormone levels. Clinical presentations included: toward female genitalia in 25 cases, male genitalia tendency in 17 cases and ambiguous external genitalia in 11 cases. Hypospadias was seen in 31 cases and short stature was seen in 8 cases. Chromosomal karyotyping of peripheral blood revealed 23 cases of sex chromosome disorders, 22 cases of 46 XY disorders, of which 3 cases were 5α-reductase deficiency and 8 cases of 46 XX disorders. Ultrasound examination showed cryptorchidism in 30 cases, including 16 cases of unilateral, 14 cases of bilateral and 1 case presenting a huge pelvic tumor. A total of 97 gonadal tissues from 53 cases of DSD were examined, including 9 cases of unilateral and 44 cases of bilateral gonads. Microscopically, 55 gonads (56.7%) showed dysplastic testes including 17 unilateral and 19 bilateral gonads. Fourteen were streak gonads (14.4%) including 8 unilateral and 3 bilateral gonadal tissues. Nine streak gonad with epithelial cord-like structures (9.3%) were found, of which 5 were unilateral and 2 were bilateral lesions. Seven gonads were ovotestis (7.2%), unilateral in 5 cases (the other side of the gonads of ovary in 4 cases, 1 case of dysplastic testes) and bilateral in 1 case. Seven gonads showed follicular-rich ovarian tissue (7.2%). One case showed bilateral dysplastic testes with gonadoblastoma and ectopic adrenal cortex. One case of streak gonad showed epithelial cord-like structures and undifferentiated glandular tissue embedded in malignant mixed germ cell tumors (mixed gonadoblastoma, dysgerminoma, mature teratoma and yolk sac tumor). One case had testicular microlithiasis. Uterus and fallopian tube structures were found in 11 cases. Immunohistochemical stains were performed in 15 cases. D2-40, PLAP and CKIT were expressed in germ cells and Calretinin, WT1 and inhibin were positive in Setoli cells. SALL4 and OCT3/4 were positive in 3 cases. Inhibin highlighted interstitial Leydig cells in 2 cases. GPC3 was positive in yolk sac tumor component. Conclusions Gonadal dysgenesis presents a broad spectrum of gonadal phenotypes with variable degrees of differentiation. The development of bilateral gonadal tissues has certain variability. Chromosomal karyotypes have no correlation with gonadal phenotypes. Accurate histopathologic diagnosis of gonadal dysgenesis plays an important role in the treatment and prognosis of the patient.

OBJECTIVETo summarize the clinicopathologic features of neuroblastic tumors (NT), and to explore the prognostic significance of MYCN amplification in NT.

METHODSThe clinicopathologic data of 267 NT were reviewed. MYCN gene amplification was detected by fluorescence in situ hybridization (FISH) in 119 cases and the relationship with pathological characteristics and prognostic significance were analyzed.

RESULTSThe study included 267 cases of children NT from patients aged from 1 day to 13 years (median 27 months). The male to female ratio was 1.43. There were 38 cases (14.2%), 43 cases (16.1%), 71 cases (26.6%), and 115 cases (43.1%) of INSS stages I, II, III and IV respectively.Favorable histology group had 157 cases (59.9%); unfavorable histology group had 110 cases (40.1%).Of the 119 NT cases with MYCN FISH performed, 18 cases (15.1%) showed amplification and the signal ratio of MYCN to CEP2 was 4.08-43.29. One hundred and one cases of non-amplified MYCN included MYCN gain in 79 cases (66.3%) and MYCN negative in 22 cases (18.5%). MYCN expression showed significant difference (P = 0.000) between ages, gender, NT type and MKI, but not INPC and clinical stage (P > 0.05).Of the 18 cases with MYCN amplification, 3 were undifferentiated, and 15 poorly differentiated; 17 had high MKI and one moderate MKI. All 18 cases were in unfavorable histology group; the overall survival rate was 3/18, with an average survival time of (17.9 ± 2.4) months.Of the 101 MYCN non-amplification cases, the overall survival rate was 68.3% (69/101), with an average survival time of (29.8 ± 1.3) months. Survival analysis showed the cases with MYCN amplification had worse prognosis (P < 0.05).

CONCLUSIONSNT were commonly diagnosed in early ages and easily to metastasize. Most of cases with favorable histology. The cases of MYCN amplification showed unfavorable histology, and the majority cases with high MKI; The patients with MYCN gene amplification had poor prognosis.

Adolescent ; Cell Differentiation ; Child ; Child, Preschool ; Female ; Gene Amplification ; Humans ; In Situ Hybridization, Fluorescence ; Infant ; Male ; N-Myc Proto-Oncogene Protein ; Neuroblastoma ; genetics ; mortality ; pathology ; Nuclear Proteins ; genetics ; Oncogene Proteins ; genetics ; Prognosis ; Survival Analysis ; Survival Rate

Background/Aims: Metabolic dysfunction–associated steatotic liver disease (MASLD) is a chronic liver disease characterized by hepatic steatosis. Ubiquitin-specific protease 29 (USP29) plays pivotal roles in hepatic ischemiareperfusion injury and hepatocellular carcinoma, but its role in MASLD remains unexplored. Therefore, the aim of this study was to reveal the effects and underlying mechanisms of USP29 in MASLD progression. Methods: USP29 expression was assessed in liver samples from MASLD patients and mice. The role and molecular mechanism of USP29 in MASLD were assessed in high-fat diet-fed and high-fat/high-cholesterol diet-fed mice and palmitic acid and oleic acid treated hepatocytes. Results: USP29 protein levels were significantly reduced in mice and humans with MASLD. Hepatic steatosis, inflammation and fibrosis were significantly exacerbated by USP29 deletion and relieved by USP29 overexpression. Mechanistically, USP29 significantly activated the expression of genes related to fatty acid β-oxidation (FAO) under metabolic stimulation, directly interacted with long-chain acyl-CoA synthase 5 (ACSL5) and repressed ACSL5 degradation by increasing ACSL5 K48-linked deubiquitination. Moreover, the effect of USP29 on hepatocyte lipid accumulation and MASLD was dependent on ACSL5. Conclusions USP29 functions as a novel negative regulator of MASLD by stabilizing ACSL5 to promote FAO. The activation of the USP29-ACSL5 axis may represent a potential therapeutic strategy for MASLD.

Background/Aims: Metabolic dysfunction–associated steatotic liver disease (MASLD) is a chronic liver disease characterized by hepatic steatosis. Ubiquitin-specific protease 29 (USP29) plays pivotal roles in hepatic ischemiareperfusion injury and hepatocellular carcinoma, but its role in MASLD remains unexplored. Therefore, the aim of this study was to reveal the effects and underlying mechanisms of USP29 in MASLD progression. Methods: USP29 expression was assessed in liver samples from MASLD patients and mice. The role and molecular mechanism of USP29 in MASLD were assessed in high-fat diet-fed and high-fat/high-cholesterol diet-fed mice and palmitic acid and oleic acid treated hepatocytes. Results: USP29 protein levels were significantly reduced in mice and humans with MASLD. Hepatic steatosis, inflammation and fibrosis were significantly exacerbated by USP29 deletion and relieved by USP29 overexpression. Mechanistically, USP29 significantly activated the expression of genes related to fatty acid β-oxidation (FAO) under metabolic stimulation, directly interacted with long-chain acyl-CoA synthase 5 (ACSL5) and repressed ACSL5 degradation by increasing ACSL5 K48-linked deubiquitination. Moreover, the effect of USP29 on hepatocyte lipid accumulation and MASLD was dependent on ACSL5. Conclusions USP29 functions as a novel negative regulator of MASLD by stabilizing ACSL5 to promote FAO. The activation of the USP29-ACSL5 axis may represent a potential therapeutic strategy for MASLD.

Objective To study the clinicopathologic features of pediatric vascular anomalies and application of ISSVA classification. Methods The clinical features, histopathologic findings and immunohistochemical results were analyzed in 117 cases of pediatric vascular anomalies encountered during the period from May 2014 to May 2015.Results A total of 117 cases of vascular anomalies were studied. The age of patients ranged from 18 hours after birth to 11 years ( mean age =34 months and median age =27 months) .There were 73 male patients and 44 female patients, with the male-to-female ratio being 1.7∶1.0.Congenital skin lesions were found in 37 cases ( 31.6%) .The common sites of involvement included head and neck region (46 cases, 39.3%), trunk (28 cases, 23.9%), extremities (14 cases, 12.0%) and internal viscera (31 cases, 26.5%).According to the new ISSVA classification, there were 74 cases of vascular malformations and 43 cases of vascular neoplasms ( ratio=1.7∶1.0 ) .The commonest vascular tumor encountered was infantile hemangioma ( 21 cases, 48.8%) , including 17 cases in proliferative phase and 4 cases in involutive phase.Thirteen cases (23.3%) of congenital hemangioma were found, with 8 cases of rapidly involuting congenital hemangioma and 5 cases of non-involutive congenital hemangioma.Three of the congenital hemangioma occurred in liver. There were 5 cases ( 11.6%) of pyogenic granuloma, 3 cases ( 7.0%) of tufted angioma and 1 case ( 2.3%) of Kaposiform hemangioendothelioma.Amongst the 74 cases of vascular malformations encountered, lymphatic malformation was found in 47 cases ( 63.5%) , venous malformation in 15 cases ( 20.2%) , lymphatic-venous malformation in 11 cases (14.9%) and arteriovenous malformation in 1 case (1.4%).All cases of vascular anomalies were all positive for CD31 on immunostaining.Glut1 and CD15 were positive both in proliferative and involutive phases of the 21 cases of infantile hemangioma, while other vascular tumors and vascular malformations were negative.Forty-seven cases of lymphatic malformation and 11 cases of lymphatic-venous malformation showed D2-40 expression.Focal positivity for D2-40 was demonstrated in 3 cases of tufted angioma and 1 case of Kaposiform hemangioendothelioma.Conclusions Vascular anomalies affecting infants and children include tumors and malformations. Accurate histopathologic diagnosis and ISSVA classification of the various types of vascular anomalies play an important role in clinical management.

Objective:To investigate the clinicopathological features of gonadal neoplastic related lesions in children with disorders of sexual development (DsD).Methods:The clinical manifestations, chromosomal karyotype, histology and immunophenotype of 12 cases of neoplastic related lesions from Guangzhou Women and Children′s Medical Center, Guangzhou were analyzed during Jan 2015 to May 2020.Results:Twelve cases of neoplastic related lesions were screened in 205 cases of DsD, and 6 patients with gonadal germ cell neoplasia aged 3-13 years with an average age of 8.3 years. There were 2 males and 4 females. Clinical features showed malformation of external genitalia in 2 cases, short stature in 2 cases, clitoral enlargement in 1 case, lower abdominal pain and a huge pelvic mass in 1 case. Chromosomal karyotyping of peripheral blood showed 2 cases of 46XY and 4 cases of 45X/46XY. Fourteen gonadal specimens were examined. Microscopically, 1 case showed dysgerminoma in left ovary, and malignant mixed germ cell tumors in right ovary, as well as gonadoblastoma (GB) and undifferentiated gonadal tissue (UGT). The remaining 5 cases were all precursor lesions of germ cell tumor. Six specimens showed GB, 3 of UGT, and 3 specimens showed germ cell neoplasia in situ (GCNIS), one of which was accompanied by intratubular seminoma and 1 was GB with GCNIS. The other 6 patients with DsD were aged from 8 months to 2 years and 5 months, including 5 males and 1 females. Clinical manifestations showed 5 cases of hypospadias and 1 case of bilateral indirect inguinal hernia. Microscopically, 6 cases showed maturation delay of gonocytes in seminiferous tubules. Immunohistochemically, the primordial germ cells/gonocytes expressed OCT3/4, PLAP and c-KIT in the 12 cases.Conclusion:Gonadal neoplasia in children with DsD is mainly precursor lesions of germ cell tumor and improved understanding of these lesions is of great significance.