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Copyright of Journal of Modern Laboratory Medicine is the property of Journal of Modern Laboratory Medicine Editorial Department and its content may not be copied or emailed to multiple sites or posted to a listserv without the copyright holder's express written permission. However, users may print, download, or email articles for individual use. This abstract may be abridged. No warranty is given about the accuracy of the copy. Users should refer to the original published version of the material for the full abstract. (Copyright applies to all Abstracts.)

Objective To investigate the independent risk factors affecting prognosis of patients with retinoblastoma (RB) and construct a nomogram to predict prognosis of patients with RB. Methods Data of 759 RB patients were collected from the SEER database. Patients were randomly assigned to the training group and validation group in a 7:3 ratio. Univariate and multivariate Cox proportional hazard regression analyses were used to determine the independent prognostic factors, based on which a nomogram was constructed. C index, calibration curve, and ROC curve were used to evaluate the predictive efficiency and calibration degree of the nomogram. Results Multivariate analysis identified independent risk factors associated with overall survival, namely, T stage and SEER stage. The C-index of SEER training set was 0.765 (95%CI: 0.744-0.786), the calibration curve was drawn, and the observed and predicted values overlapped well, indicating good consistency. The ROC curve showed that the nomogram could accurately predict three-year (AUC=0.743), five-year (AUC=0.734) and 10-year (AUC=0.720) survival rates of RB patients. Conclusion T stage and SEER stage are independent risk factors related to prognosis of RB patients, and the nomogram can accurately predict the three-year, five-year, and 10-year overall survival rates of patients.

BACKGROUND: Retinoblastoma is a common neoplasia that affects the visual organ in young children. The mortality rate is approximately 15%. In 91% of cases, surgery with enucleation is required, which significantly reduces the patients quality of life. Early diagnosis of the disease may help to correct approaches to treatment of retinoblastoma, significantly increasing the chances of preserving vision. This is important since approximately 95% of retinoblastoma cases are diagnosed before the age of 5. Using bioinformatic methods, a comprehensive analysis of the patterns and connections between the retinoblastoma-associated genes was conducted, which may further form the basis of molecular genetic testing for diagnosing this oncology. AIM: The study aimed to comprehensively analyze the genes and their products associated with retinoblastoma to reveal patterns of oncologic development. METHODS: Obtaining and sorting the list of genes using the OMIM and COSMIC databases (https://omim.org/; https://cancer.sanger.ac.uk/cosmic). Calculating gene ontology categories using DAVID and PANTHER services (https://david.ncifcrf.gov/; http://pantherdb.org/). Reconstructing the gene network using the GeneMANIA service (https://genemania.org/). Analyzing the three-dimensional (3D) structure of proteins using the PDB (RCSB) database (https://www.rcsb.org/). RESULTS: After sorting retinoblastoma-associated genes, the OMIM.org database generated a list of 139 elements. After sorting and comparison with the results of a similar query in the COSMIC database, RB1, KRAS, SYK, MYCN, and BCOR retinoblastoma-associated key genes were identified. The resulting list was analyzed for gene ontology categories using DAVID and PATHER services. The most significant categories for retinoblastoma genes were cell cycle regulators, in particular regulators of the transition from G to S phase and regulators of transcription from the RNA polymerase II promoter. Gene network structure analysis for retinoblastoma genes using the GeneMANIA service showed the existence of dense and linked gene clusters with cell cycle and transcriptional regulator genes at the center. Using the PDB database, 3D structures of key gene expression products were obtained. CONCLUSIONS: The development of molecular genetic testing of retinoblastoma for the activity of expression of associated genes and their products in the prenatal and/or postnatal period is required to improve the retinoblastoma monitoring system. The results of the study may serve as input data for this testing.

Objective To investigate the role of long non-coding RNA family with sequence similarity 83 member A antisense RNA 1 in retinoblastoma （RB） cells and its potential molecular mechanism. Methods RB cells were cultured in vitro, and Vector, pcDNA-FAM83A-AS1, NC-siRNA, and FAM83A-AS1-siRNA were transfected into cells to detect the expression of FAM83A-AS1, RB cell proliferation ability, and apoptosis rate in RB cells. Bioinformatics and dual-luciferase reporter gene experiments were performed to predict and verify the targeting relationship between FAM83A-AS1 and miR-150, and miR-150 and the high-mobility family protein A2 （HMGA 2）. The expression of miR-150 and HMGA2 mRNA in RB cells was measured by RT-qPCR； HMGA 2 protein in RB cells by Western blot； and the proliferation and apoptotic capacity of RB cells after transfection by MTT assay and flow cytometry. Results The expression of FAM83A-AS1 in the pcDNA-FAM83A-AS1 group was significantly higher than that of the Vector group and lower than that of the NC-siRNA group （P < 0.05）. The cell proliferation capacity in the pcDNA-FAM83A-AS1 group was significantly higher, and the apoptosis rate was significantly lower （P < 0.05）. Compared with NC-siRNA group, FAM83A-AS1-siRNA group showed significantly increased apoptosis rate （P < 0.05）. The miR-150 mimic significantly inhibited the proliferation capacity of RB cells and promoted cell apoptosis. However, pcDNA-HMGA 2 significantly reversed the effect of miR-150 mimic on the proliferation and apoptosis of RB cells. Conclusion Knockdown of lncRNA FAM83A-AS1 inhibited RB cell proliferation and promoted apoptosis by regulating the miR-150/HMGA2 axis. [ABSTRACT FROM AUTHOR]

Objective To investigate the independent risk factors affecting prognosis of patients with retinoblastoma (RB) and construct a nomogram to predict prognosis of patients with RB. Methods Data of 759 RB patients were collected from the SEER database. Patients were randomly assigned to the training group and validation group in a 7:3 ratio. Univariate and multivariate Cox proportional hazard regression analyses were used to determine the independent prognostic factors, based on which a nomogram was constructed. C index, calibration curve, and ROC curve were used to evaluate the predictive efficiency and calibration degree of the nomogram. Results Multivariate analysis identified independent risk factors associated with overall survival, namely, T stage and SEER stage. The C-index of SEER training set was 0.765 (95%CI: 0.744-0.786), the calibration curve was drawn, and the observed and predicted values overlapped well, indicating good consistency. The ROC curve showed that the nomogram could accurately predict three-year (AUC=0.743), five-year (AUC=0.734) and 10-year (AUC=0.720) survival rates of RB patients. Conclusion T stage and SEER stage are independent risk factors related to prognosis of RB patients, and the nomogram can accurately predict the three-year, five-year, and 10-year overall survival rates of patients. [ABSTRACT FROM AUTHOR]

Objective To investigate the effect of circROBO1 on the proliferation of retinoblastoma cells and its mechanism. Methods　 Retinoblastoma cell line Y79 was subcultured in vitro. Y79 cells in the second passage, which were in logarithmic growth phase and had good growth status, were randomly divided into the si-circROBO1 group and si-control group, which were transfected with si-circROBO1 and si-control, respectively. The cells were transfected for 6 h and continued to be cultured for 24 h. The expression of circROBO1 was detected by RT-qPCR, the cell viability was detected by CCK-8 method, and the cell proliferation was detected by cell cloning experiment. The binding sites of circROBO1 and miR-217 were predicted through the online website of CircInteractome, and the targeted regulatory relationship between the two was verified by double luciferase reporter gene test and RNA immunoprecipitation test. Results The relative expression of circROBO1 in the si-circROBO1 group was significantly lower than that in the si-control group (P< 0. 05). The cell viability and colony-forming ability of the si-circROBO1 group were significantly lower than those of the si-control group（both P<0. 05). According to the prediction of CircInteractome online website, there might be two binding sites with miR-217 in circROBO1 sequence. The double luciferase reporter gene test and RNA immunoprecipitation test showed that co-transfection of wild-type circROBO1 and miR-217 could reduce the luciferase activity, and miR-217 was mainly enriched in Y79 cells transfected with wild-type circROBO1 MS2bs vector. Conclusion CircROBO1 may promote the proliferation of retinoblastoma cells by targeting miR-217. [ABSTRACT FROM AUTHOR]

Objective To investigate the effects of overexpression of miR-515-5p on proliferation， apoptosis， and invasion of retinoblastoma cells and their mechanism of action. Methods CBX4 was predicted as the target gene of miR515-5p by bioinformatics analysis using StarBase database， which was verified by dual luciferase reporter gene assay. SORB50 cells with the most significant reduction in miR-515-5p expression among retinoblastoma cell lines SO-RB50， Y79 and HXO-RB-44 were selected for experiments. They were divided into the miR-515-5p NC group， miR-515-5p mimics group， miR-515-5p mimics+CBX4 group， and miR-515-5p mimics+CBX4-NC group； miR-515-5p NC， miR-515-5p mimics， miR-515-5p mimics+CBX4 overexpression vector， and miR-515-5p mimics+CBX4 empty vector were transfected， respectively. The expression of miR-515-5p and CBX4 mRNA in each group was detected by real-time fluorescent quantitative PCR， and the expression of CBX4 protein was detected by Western blotting. The cell proliferation ability （OD value） was detected by CCK-8 at 12，24，48， and 72 h after transfection. The apoptosis was detected by TUNEL experiment. Transwell chamber assay was used to detect cell invasion ability. Results The relative expression of miR-515-5p in the miR-515-5p mimics group， miR-515-5p mimics+CBX4 group， and miR-515-5p mimics+CBX4-NC group was significantly higher than that in miR-515-5p NC group （all P<0. 05）. The relative expression levels of CBX4 mRNA and protein in the miR-515-5p mimics group， miR-515-5p mimics+CBX4-NC group， and miR-515-5p mimics+CBX4 group were lower than those in the miR-515-5p NC group at 48 and 72 h； the decreased in the miR-515-5p mimics group and miR-515-5p mimics+CBX4-NC group was more significant （all P<0. 05）. At 12 of culture， no significant difference was found in the OD value of cells among groups （P>0. 05）. At 24 h of culture， the OD values of the miR-515-5p mimics group and miR-515- 5p mimics+CBX4-NC group were lower than those of miR-515-5p NC group （all P<0. 05）. Compared with the miR-515-5p NC group and miR-515-5p mimics+CBX4 group， the apoptosis rate increased and the number of invasive cells decreased in the miR-515-5p mimics group and miR-515-5p mimics+CBX4-NC group （all P<0. 05）. Conclusions Overexpression of miR-515-5p can reduce the proliferation and invasion of SO-RB50 cells in retinoblastoma cells， and promote their apoptosis， which may be related to the inhibition of CBX4 expression. [ABSTRACT FROM AUTHOR]

Objective To explore the effect of lncRNA ADPGK-AS1 on the proliferation and apoptosis of retinoblastoma cells and its possible mechanism. Methods The tumor tissues of 31 patients with retinoblastoma admitted to Henan Provincial Eye Hospital from February to June 2020 and their corresponding normal tissues adjacent to the cancer were collected. The expression levels of lncRNA ADPGK-AS1 and miR-200b-5p in retinoblastoma tissues and normal adjacent tissues were detected by real-time fluorescence quantitative polymerase chain reaction (qRT-PCR). Human retinal epithelial cell ARPE-19, human retinoblastoma cell Y-79 and WERI-Rb-1 were cultured in vitro. The expression levels of lncRNA ADPGK-AS1 and miR-200b-5p were detected by qRT-PCR. Y-79 cells were randomly divided into si-con group, si-lncRNA ADPGK-AS1 group, miR con group, miR-200b-5p group, si-lncRNA ADPGK-AS1+ anti-miR con group, and si-lncRNA ADPGK-AS1+ anti-miR-200b-5p group. The proliferation, cloning and apoptosis of cells in each group were detected by tetramethylazol blue method, plate cloning test and flow cytometry, respectively. The targeting relationship between lncRNA ADPGK-AS1 and miR-200b-5p was detected by double luciferase report test, and the expression level of cleaved-caspase-3 protein was detected by western blot. Results Compared with the adjacent tissues, the expression of lncRNA ADPGK-AS1 in retinoblastoma tissues was increased (P<0.05), while the expression of miR-200b-5p was decreased (P<0.05). Compared with ARPE-19 cells, the expression of lncRNA ADPGK-AS1 in Y-79 and WERI-Rb-1 cells was increased (P<0.05), while the expression of miR-200b-5p was decreased (P<0.05). Compared with the si-con group, the cell viability of the si-lncRNA ADPGK-AS1 group was reduced (1.06±0.09 vs 0.53±0.05, P<0.05), the number of cell clone formation was reduced (114.00±8.03 vs 57.00±4.13, P<0.05), while the apoptosis rate [(7.93±0.68)% vs (25.43±1.94)%] and the protein level of cleaved-caspase-3 were increased (P<0.05). Compared with the miR-con group, the cell viability of the miR-200b-5p group was decreased (1.05±0.08 vs 0.57±0.05, P<0.05), the number of cell clone formation was decreased (118.00±10.02 vs 64.00±5.13, P<0.05), while the apoptosis rate [(7.89±0.71)% vs (23.15±1.62)%] and the protein level of cleaved-caspase-3 were increased (P<0.05). lncRNA ADPGK-AS1 could target the expression of miR-200b-5p. Compared with the si-lncRNA ADPGK-AS1+ anti-miR-con group, cell viability of the si-lncRNA ADPGK-AS1+ anti-miR-200b-5p group was increased (0.53±0.04 vs 1.25±0.10, P<0.05), and the number of cell clones was increased (54.00±4.39 vs 125.00±10.03, P<0.05), while the rate of apoptosis [(25.38±1.53)% vs (9.76±0.71)%] and the protein level of cleaved-caspase-3 were decreased (P<0.05). Conclusion Interfering with the expression of lncRNA ADPGK-AS1 could inhibit the proliferation and clone formation and induce apoptosis of retinoblastoma cells by targeting the expression of miR-200b-5p. [ABSTRACT FROM AUTHOR]

The patient is a 2-year-old male. The family consulted the Department of Ophthalmology, Shanghai Ninth People's Hospital, after noticing a white reflection in the pupil area of the child's right eye for 6 days. Following a thorough ocular and systemic examination, the patient was diagnosed with retinoblastoma (Group E, cT2bN0M0) of the right eye. The right eye was enucleated and classified as pathological stage pT3cN0M0. Postoperatively, systemic intravenous chemotherapy with the VEC regimen was administered. Genetic testing revealed a germline mutation in the RB1 gene: c.874 (exon9) delT (p.Tyr292fsTer9), necessitating close monitoring of the socket during follow-up visits. Three months after the operation, fundus examination revealed yellow-white lesions in the left eye, and bilateral retinoblastoma was diagnosed (Group E in the right eye, Group C in the left eye). Based on the ICRB and pTNM stages, the patient underwent six rounds of systemic intravenous chemotherapy and three rounds of cryotherapy in the left eye. No recurrence was detected with a 4-year follow-up. The patient was initially diagnosed with unilateral retinoblastoma, but later developed the disease in the contralateral eye during treatment, which was a case of metachronous bilateral retinoblastoma.

Copyright of Journal of Modern Laboratory Medicine is the property of Journal of Modern Laboratory Medicine Editorial Department and its content may not be copied or emailed to multiple sites or posted to a listserv without the copyright holder's express written permission. However, users may print, download, or email articles for individual use. This abstract may be abridged. No warranty is given about the accuracy of the copy. Users should refer to the original published version of the material for the full abstract. (Copyright applies to all Abstracts.)

Copyright of Journal of Modern Laboratory Medicine is the property of Journal of Modern Laboratory Medicine Editorial Department and its content may not be copied or emailed to multiple sites or posted to a listserv without the copyright holder's express written permission. However, users may print, download, or email articles for individual use. This abstract may be abridged. No warranty is given about the accuracy of the copy. Users should refer to the original published version of the material for the full abstract. (Copyright applies to all Abstracts.)

Retinoblastoma (RB) seriously damages children’s vision health and life safety. This study established a new method of optical coherence tomography (OCT) assisted RB zebrafish model construction, and used OCT to evaluate the distribution and survival status of RB cells in the eyes in vivo. Twenty-four normal zebrafish were used in this study, of which 21 were used to construct the RB zebrafish model and the remaining 3 were used as control. The specific method was to inject fluorescent RB cell into the eyeball through an injection under OCT real-time imaging. The distribution and survival status of cell were observed at the 0th, 24th, 48th, and 72th hour after cell injection. At the same time, the zebrafish eyes were imaged with a stereo fluorescent microscope (SFM) before and after modeling, as a control. The results showed that high-resolution, large-depth threedimensional distribution of RB cell clusters in the eyes could be obtained by OCT. Further OCT image intensity analysis could quantify the number of RB cells, and the results were highly matched with the results of the SFM images. In conclusion, the strategy of combining zebrafish model with OCT imaging technology is expected to provide new methods and equipment for the study of RB. [ABSTRACT FROM AUTHOR]

Objective To analyze the characteristics of gene mutations and clinical phenotypes in children with retinoblastoma. Methods We performed molecular genetic screening in 23 children with sporadic retinoblastoma (bilateral in 4 and unilateral in 19 children) treated in 2018 to 2019 in the Second Affiliated Hospital of Chongqing Medical University by whole exon sequencing and methylation sequencing of the peripheral blood samples. The sequence data were analyzed in relation with the clinical features of the patients to explore the correlation between the genotypes and clinical phenotypes. Results Five pathological germline mutations of RB1 gene were identified in the 23 cases, including 3 cases with point mutations in RB1 gene (nonsense mutation c.1333C>T, missense mutation c.380G>A, and frameshift mutation c.1940delT), 1 case with heterozygous deletion of exon 5-17 in RB1 gene and 1 with heterozygous deletion of about 14 Mb fragment of RB1 gene and its adjacent sequence. The patients with genetic mutation had an average onset age of 5.4 months, significantly younger than the others (average onset age of 20.2 months). and a significant correlation was found between gene mutations and the age of disease onset (P < 0.01, r=0.592). The detection rate of gene mutations was significantly higher in cases of bilateral tumors than in those with unilateral tumor (3/4 vs 2/19; Chi-square=8.074, P < 0.01). Conclusion We identified for the first time 2 new germline copy number variants of RB1 gene in children with retinoblastoma, which expand the spectrum of RB1 gene variations. Germline variation of RB1 gene is associated with a higher likelihood of bilateral eye involvement and an earlier onset of retinoblastoma in children.

Objective To investigate the characteristics of clinical treatment and prognosis of 118 cases of retinoblastoma (RB). Methods A total of 118 RB children treated in our department from February 2008 to February 2018 were recruited in this study. All of them were evaluated by the International Classification for Intraocular Retinoblastoma (IIRC, A~E stage) and extraocular staging standard. Mainly according to their tumor stage, combined with their physical condition and the wishes of their parents, treatments such as intravenous chemotherapy, transpupillary thermotherapy (TTT), freezing and enucleation were applied. All cases were followed up for 1 to 10 years, and the clinical efficacy and prognosis of the patients were analyzed retrospectively. Results The 118 enrolled children (154 eyes) were 70 males and 48 females, at an average age of 25.94±20.88 months (1~122 months) at first visit, and 36 of them suffered from bilateral RB and 82 from single. Among them, 41 received chemotherapy, 57 chemotherapy combined with TTT, 10 chemotherapy combined with freezing, and 10 enucleation only. After treatment, 76 eyes (76/154) were salvaged, with a total salvage rate of 49.35%, and of them, 47 eyes were treated with chemotherapy combined with TTT, accounting for 61.84% (47/76). Seventy-eight eyes were enucleated, with a total extraction rate of 50.65% (78/154), and among 63 eyes at stage E, 53 were enucleated, accounting for 84.13%. During the mean follow-up time of 86.66±5.69 months, 22 (18.64%) children died, and 96 cases (81.36%) survived. The 5-year cumulative survival rate of single eye was 71.40%, and of bilateral was 80.00%. Conclusion Intravenous chemotherapy combined with TTT is one of the main ways for RB salvage. Enucleation is still the primary treatment for stage E eyes.

Regulating the process of epithelial-mesenchymal transition(EMT) is an essential strategy to inhibit tumor growth and metastasis. This study is based on the EMT process of retinoblastoma and constructs quercetin(QUE) and doxorubicin(DOX) co-loaded liposome(QD Lipo) to investigate the therapeutic effect and mechanisms of combined QUE and DOX treatment on retinoblastoma. Single-factor experiments were conducted to optimize the prescription process of QD Lipo. Eventually, spherical particles with a diameter of(108.87±1.93) nm, a PDI of 0.13±0.02, and a Zeta potential of(-34.83±1.92) mV were obtained. The encapsulation rates of QUE and DOX were 96.20%±4.40% and 91.17%±4.41%, respectively. Y79 human retinoblastoma cells were used as an in vitro cellular model, and confocal microscopy demonstrated that QD Lipo could enhance Y79 uptake efficiency. The CCK-8 assay confirmed that the optimal combination therapy effect of QUE and DOX occurred at a mass ratio of 1∶1 to 1∶2. Flow cytometry showed that QD Lipo enhanced the induction of apoptosis in Y79 cells. Western blot analysis revealed that QD Lipo significantly reduced the expression of EMT pathway-related proteins vimentin and α-SMA. Fluorescence assays detected a significant decrease in ROS levels in Y79 cells after treatment with QD. These results indicated that liposomal co-delivery of QUE and DOX can enhance drug delivery efficiency to retinoblastoma cells, inhibit the EMT process in retinoblastoma by downregulating ROS levels, and enhance the cytotoxicity of DOX against retinoblastoma.

Retinoblastoma is the malignant intraocular tumor most commonly happened on children, especially the children under 3 years old. It may lead to blindness, eyeball removal and even death, which severely threaten the safety and quality of children's life. Exploring the origin of tumor cells is conducive to the reveal of the pathogenesis as well as the guidance on diagnosis, classification and treatment of retinoblastoma. Based on the studies on retinal development, three theories regarding the cell origin of retinoblastoma, namely the origin of retinal precursor cells, the origin of cone cells and the origin of horizontal cells or Müller glia cells have lately been proposed. This article reviews and summarizes the research of the origin cells in retinoblastoma. [ABSTRACT FROM AUTHOR]

Objective To investigate the role of autophagy in cisplatin resistance of retinoblastoma Y79 cells and its mechanism. Methods The IC50 of cells was detected by CCK-8 assay. Y79 cells were randomly divided into control group, Cisplatin group and cisplatin combined with autophagyroup). Western blot, cell autophagy staining kit (MDC method) and transmi inhibitor 3-methyladenine group(Cis+3-MA gssion electron microscopy were used to observe cell autophagy. CCK-8 assay was used to detect the inhibitory rate of cisplatin on Y79 cells. Annexin V/PI double staining was used to detect cells apoptosis. q-PCR was applied to detect drug-resistance-related genes transcription levels. The expression of CaMKK2, p-AMPK, mTORC1 and LC3Ⅱ were detected by Fluo-4 AM calcium ion fluorescent probe staining. Results Autophagy of retinoblastoma Y79 cells was induced by cisplatin. Autophagy inhibitor 3-MA reduced the level of autophagy. Compared with cisplatin group, the cisplatin inhibition rate was increased, the apoptosis rate was increased and the drug-resistance-related gene transcription level was down-regulated in Cis+3-MA group. After adding cisplatin, the level of intracellular calcium was increased, the expressions of CaMKK2, p-AMPK and LC3Ⅱ were up-regulated and the expression of mTORC1 was down-regulated. Conclusion Cisplatin-induced autophagy is protective for the drug resistance of retinoblastoma Y79 cells and the inhibition of autophagy could improve tumor cells resistance to cisplatin. Cisplatin could induce autophagy in Y79 cells by Ca2+ activation of CAMKK2/AMPK/mTORC1 pathway.

Objective • To investigate the safety of treatment with ophthalmic artery cannulation for intra-arterial chemotherapy (IAC) in children with retinoblastoma (RB) during general anesthesia. Methods • A total of 60 children with RB who underwent ocular artery interventional chemotherapy under general anesthesia in Shanghai Ninth People's Hospital, Shanghai Jiao Tong University School of Medicine from September 2015 to August 2018 were collected. Induction of anesthesia was performed with rapid induction of endotracheal intubation with midazolam, fentanyl, propofol and rocuronium. Sevoflurane and oxygen\were administered for maintenance of general anesthesia, with intermittent injection of rocuronium and fentanyl. Intraoperative continuous monitoring oXhemodynamic parameters, respiratory parameters (EtC02, oxygen saturation and inspiratory peak pressure) and sevoflurane minimum alveolar concentration (MAC) was performed, and intraoperative pulmonary compliance, hypoxemia, hypotension and other cardiopulmonary adverse events were observecKand recorded. Results • Over a 3-year period, 185 treatment sessions were performed in 60 patients. Thirty-two cardiopulmonary adverse events were observed in 20 patients, and the incidence rate was 17.2%, mainly including severe decrease in lung compliance, hypoxemia and arterial hypotension. All severe decreases in lung compliance occurred within 1 or 2 minutes after catheter insertion in the ophthalmic artery. After active treatment with propofol, phenylephrine and epinephrine, no death and permanent sequelae occured. Conclusion • An appreciable incidence of trigeminocardiac reflex to intra-ophthalmic artery infusion of chemotherapy in patients with RB is found. Both intcrvcntionalists and anesthesiologists should be aware of this potential event and be prepared to provide immediate resuscitative measures. [ABSTRACT FROM AUTHOR]

Objective: To investigate the effect ofhypoxia on the invasion ability ofretinoblastoma cell and its underlying mechanism. Methods: HXO-RB44 cells were cultured under normoxia (21%O O2) and hypoxia(1%O O2) conditions for 24 hours. The effect of hypoxia on invasion ability of HXO-RB44 cells were monitored with transwell invasion assay; the mRNA and protein expression levels of HIF-1a and MMP9 were detected by RT-PCR and western blot; luciferase assay was performed to assess the HIF-1a regulation by hypoxia, and MMP9 regulation by HIF-1a. Furthermore, HIF-1a and MMP9 siRNA was used to investigate the effect of HIF-1a/MMP9 signaling on hypoxia induced cell invasion. Results: The invaded cell number，mRNA and protein expression of HIF-1a and MMP9 in hypoxia group were higher than those in the normoxia group (P<0.05). The activity of MMP9 in HIF-1a overexpression group was higher than that of the vector group (P<0.05). Compared with the normoxia group，the activity ofHIF-1a in hypoxia group was significantly elevated (P< 0.05). Compared with negative control group，the mRNA and protein expression of HIF-1a and MMP9 in HIF-1a siRNA group were re¬markedly decreased (P<0.05)，whereas the mRNA and protein expression of HIF-1a in MMP9 siRNA group didn't change. The invaded cells in HIF-1a siRNA group and MMP9 siRNA group were significantly decreased (P<0.05). Conclusion: Hypoxia promotes the inva¬sion ofretinoblastoma cell line HXO-RB44 and the activation ofHIF-1a/MMP9 signaling pathway is involved in this process. [ABSTRACT FROM AUTHOR]

Objective: To study the expression of regenerating islet-derived family member 4 (RegⅣ) in retinoblastoma and its correlation with angiogenesis. Methods: 16 cases of patients with retinoblastoma who underwent operation in our hospital from June 2012 to June 2014 were selected as the research object.The positive expression rates of RegⅣ, VEGF-A and VEGF-C between retinoblastoma tissues and normal tissues were compared, the correlation of clinicopathological features with the expressions of RegⅣ, VEGF-A and VEGF-C were analyzed, and theh microvessel density (MVD) between RegⅣ positive expression tumor tissue and normal tissue were compared. Results: Compared with the normal tissues, the Reg Ⅳ, VEGF-A and VEGF-C expressions were higher in the retinoblastoma tissues (P＜0.05). The Reg Ⅳ,VEGF-A, VEGF-C expressions were significantly correlated with the clinical stage,pathological type,optic nerve invasion retinoblastoma (P＜0.05), but had nothing to do with gender, site of onset (P＞0.05). Pearson correlation analysis showed the expression of Reg Ⅳ was positively correlated with VEGF-A(r=0.696,P＜0.05); the expression of Reg Ⅳ was positively correlated with VEGF-C(r=0.728,P＜0.05). The MVD value of Reg Ⅳ positive expression was (59.67± 1.43), which was significantly higher than that of the normal retinal tissues (30.53± 2.31) (P＜0.05); the MVD value of Reg Ⅳ negative expression tissues was (38.04± 2.86), which had no difference compared with the normal retinal tissues (P＞0.05). Conclusions: The positive expression rates of Reg Ⅳ was highly expressed in the retinoblastoma tissue and correlated with the invasion and migration, it could accelerate the development of the retina through promoting the angiogenesis mediated by VEGF-A and VEGF-C. [ABSTRACT FROM AUTHOR]

Objective: To explore the effect of lncRNA ADPGK-AS1 on the proliferation and apoptosis of retinoblastoma cells and its possible mechanism. Methods: The tumor tissues of 31 patients with retinoblastoma admitted to Henan Provincial Eye Hospital from February to June 2020 and their corresponding normal tissues adjacent to the cancer were collected. The expression levels of lncRNA ADPGK-AS1 and miR-200b-5p in retinoblastoma tissues and normal adjacent tissues were detected by real-time fluorescence quantitative polymerase chain reaction (qRT-PCR). Human retinal epithelial cell ARPE-19, human retinoblastoma cell Y-79 and WERI-Rb-1 were cultured in vitro . The expression levels of lncRNA ADPGK-AS1 and miR-200b-5p were detected by qRT-PCR. Y-79 cells were randomly divided into si-con group, si-lncRNA ADPGK-AS1 group, miR con group, miR-200b-5p group, si-lncRNA ADPGK-AS1+ anti-miR con group, and si-lncRNA ADPGK-AS1+ anti-miR-200b-5p group. The proliferation, cloning and apoptosis of cells in each group were detected by tetramethylazol blue method, plate cloning test and flow cytometry, respectively. The targeting relationship between lncRNA ADPGK-AS1 and miR-200b-5p was detected by double luciferase report test, and the expression level of cleaved-caspase-3 protein was detected by western blot. Results: Compared with the adjacent tissues, the expression of lncRNA ADPGK-AS1 in retinoblastoma tissues was increased ( P <0.05), while the expression of miR-200b-5p was decreased ( P <0.05). Compared with ARPE-19 cells, the expression of lncRNA ADPGK-AS1 in Y-79 and WERI-Rb-1 cells was increased ( P <0.05), while the expression of miR-200b-5p was decreased ( P <0.05). Compared with the si-con group, the cell viability of the si-lncRNA ADPGK-AS1 group was reduced (1.06±0.09 vs 0.53±0.05, P <0.05), the number of cell clone formation was reduced (114.00±8.03 vs 57.00±4.13, P <0.05), while the apoptosis rate [(7.93±0.68)% vs (25.43±1.94)%] and the protein level of cleaved-caspase-3 were increased ( P <0.05). Compared with the miR-con group, the cell viability of the miR-200b-5p group was decreased (1.05±0.08 vs 0.57±0.05, P <0.05), the number of cell clone formation was decreased (118.00±10.02 vs 64.00±5.13, P <0.05), while the apoptosis rate [(7.89±0.71)% vs (23.15±1.62)%] and the protein level of cleaved-caspase-3 were increased ( P <0.05). lncRNA ADPGK-AS1 could target the expression of miR-200b-5p. Compared with the si-lncRNA ADPGK-AS1+ anti-miR-con group, cell viability of the si-lncRNA ADPGK-AS1+ anti-miR-200b-5p group was increased (0.53±0.04 vs 1.25±0.10, P <0.05), and the number of cell clones was increased (54.00±4.39 vs 125.00±10.03, P <0.05), while the rate of apoptosis [(25.38±1.53)% vs (9.76±0.71)%] and the protein level of cleaved-caspase-3 were decreased ( P <0.05). Conclusion: Interfering with the expression of lncRNA ADPGK-AS1 could inhibit the proliferation and clone formation and induce apoptosis of retinoblastoma cells by targeting the expression of miR-200b-5p.

Retinoblastoma (RB) is the most common intraocular malignant tumor in children. Chemotherapy remains the main method for the treatment of intraocular RB in term of saving affected eyes. Intra-arterial chemotherapy is a novel way to treat intraocular RB and can avoid systematic toxic side effects caused by intravenous chemotherapy. This article reviews the research advances on intra-arterial chemotherapy for the treatment of RB. [ABSTRACT FROM AUTHOR]

Retinoblastoma (RB) is the most common intraocular malignant tumor in infants and young children. The key causative factors in the progression of RB remain unclear. Therefore, identifying genes closely associated with RB progression may provide important clues for disease diagnosis and gene therapy. However, tumor tissues have strong cellular heterogeneity. There may be significant differences in cell function and gene expression among cells in different pathological states. In this study, we downloaded single-cell transcriptome sequencing data of RB tumors and adjacent tissues from the GEO public database. Subsequently, we analyzed RB tumor transcriptional profiles with different disease duration at the single-cell level and identified cell groups and gene sets potentially associated with RB progression. The results showed that the tumor tissue and the adjacent tissues had overall consistency in the single-cell transcriptional map, but there were obvious differences in the distribution proportions of G1 phase cells, G2 phase cells, and microglia cells of cone precursors in RB tumor and the adjacent tissues. Furthermore, the role of three cell populations in the progression of RB tumors was emphatically analyzed. We found that in the early stage of RB tumors, cone precursor cells proliferated abnormally in G1 phase. With the progression of RB tumors, the proportion of cone precursor cells in G2 phase increased significantly. Meanwhile, the results of differential analysis of microglial populations during RB progression showed that the key genes mainly involved in immune response include RPL23 , B2M , and HLA superfamily genes. This study provides new perspectives and data resources for the research of RB pathogenesis and progress.

Objective To investigate the effects of diallyl disulfide (DADS) on proliferation and invasion of retinoblastoma cells Y79 and to investigate the mechanism. Methods The retinoblastoma cells Y79 were cultured and divided into the miRNA negative control group (tansfected by 40 μmol/L scramble), miR-222 group (tansfected by 40 μmol/L miR-222-mimics), DADS negative control group (added with 10 μmol/L DMSO) and DADS group (added with 200μmol /L DADS) and DADS + miR-222 I group (transfected by 40 μmol /L miR-200b-inhibitors + 200 μmol /L DADS). The expression of miR-222 treated by different concentrations of DADS in Y79 cells was detected by qRT-PCR. The proliferation and invasion of retinoblastoma cells Y79 in vitro were determined with MTT and Transwell invasion assay. Results The miR-222 expression of Y79 cells treated with 0,25,50,100,200 and 400 μmol/L DADS was 2. 823 ± 0. 250,2. 343 ± 0. 226,1. 955 ± 0. 267,1. 567 ± 0. 096,0. 875 ± 0. 189 and 0. 718 ± 0. 126, respectively. The OD570 of miR-222 was respectively 0. 727 ± 0. 167,0. 949 ± 0. 070,0. 712 ± 0. 178,0. 497 ± 0. 126 and 0. 351 ± 0. 102 in the miRNA negative control group,miR-222 M group, DADS negative control group, DADS group and DADS + miR-222 I group. Transwell invasion assay results showed that the number of cells permeating the matrigel significantly decreased to 128 ± 8,179 ± 16,127 ± 12,76 ± 8 and 45 ± 14. Significant difference was respectively found between the miR-222 M group,DADS group,DADS + miR-222 I group and the miRNA negative control group and DADS negative control group( all P <0. 05). Conclusion DADS can inhibit the proliferation and invasion of retinoblastoma cells Y79 by down-regulating the expression of miR-222. [ABSTRACT FROM AUTHOR]

Objective To investigate the value of the clinical application of multislice spiral CT in the measurement of orbital volume. Methods 12 cases of congenital microphthalmia or ocular dysplasia, 4 cases of traumatic eyeball atrophy in children, 15 cases retinoblastoma enucleation, and 64 cases of orbital blowout fracture were plain scanned by CT Measuring software, and the orbital volumes of both side of sick and healthy eyes were measured. Results The volumes of 23 cases of congenital microphthalmia or ocular dysplasia, traumatic eyeball atrophy in children or retinoblastoma enucleation were less than the healthy eyes. The volumes of eyes suffered from orbital blowout fracture were greater than the healthy eyes. Conclusion The technology can evaluate the degree of development of children eyes and estimate the influence of the therapy by the orbital implants. Meanwhile, it can measure the volume of orbital blowout fracture and predict the enophthalmos situation. It is significantly helpful to the therapeutic planning and the assessment in pre-and post-operative. [ABSTRACT FROM AUTHOR]

Objective: To analyze the clinical features and fundus performance of patients with retinocytoma. Methods: Retrospective case series study. Ten patients (12 eyes) diagnosed with retinocytoma were enrolled in Department of Ophthalmology of Eye & ENT Hospital of Fudan University from January 2006 to December 2019. Clinical data were reviewed, including gender, age, initial symptoms, tumor size and location, fundus manifestations and regression patterns. Results: There were 7 males and 3 females aged from 5 to 39 years, with a median age of 14 years. Eight patients had unilateral retinocytoma, while 2 patients had bilateral retinocytoma. The most common complaints were strabismus (4 patients) and blurred vision (3 patients). The average maximum base diameter of the tumor was 5.70 mm (2.69 to 8.69 mm), and the average thickness was 2.28 mm (0.61 to 3.76 mm). Tumors in 12 eyes were all of a single focus, 5 in the posterior pole, 5 in the midperiphery and 2 in the periphery. Ophthalmoscopic features of retinocytoma included a translucent retinal mass in 10 eyes, a calcification mass in 11 eyes, with well-defined retinal pigment epithelial alteration in 12 eyes and a zone of chorioretinal atrophy in 6 eyes. The regression patterns were type 1 in 2 eyes, type 2 in 1 eye and type 3 in 9 eyes. The gene test showed RB1 mutation in 1 patient. During the average follow-up of 28.7 months, all of the tumors were stable without malignant transformation. Conclusions: Retinocytoma is a rare benign retinal tumor with characteristic fundus manifestations. Patients with retinocytoma should be monitored closely. (Chin J Ophthalmol, 2021, 57: 526-530) .

Objective: To investigate whether DNMT1 protein induces retinoblastoma proliferation by silencing MEG3 gene., Methods: Two retinoblastoma cell lines (HXO-RB44 and SO-RB50) and a normal human retinal pigment epithelial (RPE) cell line were transfected with the plasmid pcDNA-DNMT1 or si-DNMT1 for up-regulating or interference of DNMT1 expression, and with pcDNA-MEG3 or si-MEG3 for up-regulating or interference of MEG3 expression. Western blotting was used to detect the changes in the expression of DNMT1 protein in the transfected cells, and CCK-8 and EdU assays were used to detect the changes in cell proliferation. Real-time quantitative PCR (qRT-PCR) was performed to detect MEG3 expression in SO-RB50 and HXO-RB44 cells after transfection, and the methylation level of MEG3 gene promoter after interference of DNMT1 expression was detected using methylation-specific PCR., Results: SO-RB50 and HXO-RB44 cells showed significantly increased expression of DNMT1 protein as compared with normal RPE cells ( P < 0.05). In HXO-RB44 cells, transfection with pcDNADNMT1 resulted in significantly increased expression of DNMT1 protein, enhanced cell proliferation ability, and significantly reduced expression of MEG3 ( P < 0.05). In SO-RB50 cells, transfection with si-DNMT1 significantly reduced the expression of DNMT1 protein, suppressed the cell proliferation, and increased MEG3 expression ( P < 0.05). Interference of DNMT1 significantly reduced the methylation level of MEG3 gene promoter. After reversing the regulatory effect of DNMT1 on MEG3 gene, DNMT1 protein showed significantly weakened ability to regulate retinoblastoma cell proliferation ( P < 0.05)., Conclusions: In retinoblastoma cells, the up-regulation of DNMT1 protein induces promoter methylation and inactivation of MEG3 gene and eventually leads to abnormal cell proliferation.

Objective: To evaluate the potential association between optic nerve invasion and optic nerve obscuration during treatment of advanced retinoblastoma. Methods: Retrospective case series study. Medical records of 77 patients (77 eyes) with advanced retinoblastoma (Group D/E) who were treated with primary or secondary enucleation in the Ophthalmology Department of Peking University People's Hospital from January 1 st 2012 to December 31 th 2015 were retrospectively reviewed. RetCam photographs under general anesthesia at diagnosis and each subsequent follow-up were evaluated for complete obscuration of the optic nerve. The primary endpoints included prelaminar invasion, postlaminar invasion and optic nerve transection invasion. Group difference was calculated with chi-square. Results: There were 46 boys and 31 girls in the study. The mean age at the first diagnosis was (27.1±22.1) months. The optic nerve was obscured in 62 eyes (80.5%) at the first diagnosis and 61 eyes (79.2%) at the last ocular examination prior to enucleation. Twenty-nine eyes (37.7%) underwent primary enucleation. Forty-eight eyes (62.3%) were treated with eye-preserving therapy, followed by enucleation. Fourteen eyes (18.2%) were in Group D and 63 eyes (81.8%) were in Group E. Histopathologic analysis of enucleated eyes without optic nerve obscuration (16 eyes) showed prelaminar invasion in 7 eyes, postlaminar invasion in 2 eyes and optic nerve transection invasion in 0 eyes. Histopathologic analysis of enucleated eyes with optic nerve obscuration (61 eyes) showed prelaminar invasion in 26 eyes, postlaminar invasion in 9 eyes and optic nerve transection invasion in 4 eyes. The difference between two groups did not achieve statistical significance ( P =0.935, 1.000, 0.296). Histopathologic analysis of enucleated eyes with persistent complete obscuration of the optic nerve showed a high risk factor in 10 eyes (10/40), while in 1 eye (1/8) the optic nerve was visible at the initial presentation and obscured before secondary enucleation ( P =0.529). Conclusion: Optic nerve obscuration at the last examination prior to enucleation may not be associated with postlaminar optic nerve invasion in advanced retinoblastoma. (Chin J Ophthalmol, 2020, 56: 681-687) .

Retinoblastoma, the most frequent malignant intraocular tumor in childhood, is caused by oncogenic mutations in the RB1 tumor suppressor gene. Identification of these mutations in patients is important for genetic counseling and clinical management of relatives at risk, and thus probands are conventionally applied gene detection in developed countries. However, gene diagnosis is still in the elementary period in China. This article reviews the characteristics of retinoblastoma genetics and the current status of genetic testing in China, so as to attract more attention from ophthalmologists and to promote regulated gene diagnosis in clinical work. Not only does good understanding of retinoblastoma genetics support optimal care for retinoblastoma children and their families, but also promotes the development in foundational research. (Chin J Ophthalmol, 2019, 55: 806-810) .

Objective: To analyze the histopathological features and histopathological risk factors (HRF) of tumor recurrence after vitrectomy in patients with retinoblastoma (RB). Methods: Retrospective case series. Twenty-nine patients (29 eyes) who underwent enucleation from April 2014 to April 2019 at the Beijing Institute of Ophthalmology, Beijing Tongren Eye Center, Beijing Tongren Hospital, Capital Medical University due to tumor recurrence after vitrectomy in the other hospitals were enrolled and archived. The pathological sections of the other hospitals were read by the pathologists of the department and the pathological report was written. Histopathological features and HRF were analyzed in this group of patients. The HRF used in this study was defined as tumor invasion of the optic nerve, the choroid (the largest diameter ≥ 3 mm and mostly reaching the inner sclera), the sclera, the anterior segment of the eye, and tumor passing through the scleral and growing outside the eyeball. Results: Of the 29 patients with RB recurrence after vitrectomy, 18 (62.1%) were male and 11 (37.9%) were female. The age was 1-10 years with a median age of 3 years. Among 29 eyes, the tumor invaded the ciliary body in 19 eyes (65.5%), the iris in 14 eyes (48.3%), the anterior chamber in 10 eyes (34.5%), and the anterior chamber angle in 7 eyes (24.1%). Of the 29 eyes, 27 eyes (93.1%) had HRF. The most common HRF was invading the anterior segment of the eye(77.8%, 21/27). Conclusions: In patients with recurrence after vitrectomy for RB, the most common tumor invasion site in histopathology is the anterior segment of the eye, particularly the ciliary body. More than 90% of the patients have HRF. Vitrectomy for RB treatment requires a rigorous surgical indication assessment. ( Chin J Ophthalmol, 2019, 55: 854-859 ).

Objective: To investigate the role of Janus kinase (JAK) inhibitor AG490 in the anti-proliferation and cell cycle in human retinoblastoma HXO-RB44 cell lines in vitro, and to explore its effect on the expression of JAK2/signal transducer and activator of transcription 3 (STAT3).
 Methods: Cells were divided into an experiment group and a control group, and the experiment group was further divided into 6 sub-groups according to different AG490 concentrations (6.25, 12.50, 25.00, 50.00 or 100.00 μmol/L). Cell proliferation in the different groups was analyzed by cell vitality determination. Cell cycle distribution and apoptosis rate were examined by flow cytometry. The protein levels of STAT3, p-STAT3 and vascular endothelial growth factor (VEGF) were detected by Western blot.
 Results: After 48 h treatment with AG490, the viability of HXO-RB44 cells was reduced in a concentration-dependent manner. Compared with the control group, there was no significant difference in the experiment groups except the 6.25 μmol/L group (all P>0.05). The apoptosis rates in the experiment groups were significantly increased with increase in concentration of AG490 compared with that in the control group (all P<0.05). The cell ratio in the G1 phase in 50 or 100 μmol/L group was increased, whereas the cell ratio in the S phase was decreased. Western blot results showed that the expressions of STAT3 and p-STAT3 in the experiment groups were dramatically reduced with the increase in concentration of AG490 compared with that in the control group (all P<0.05). VEGF expression didn't obviously change in the experiment groups with AG490 concentration less than 12.5 μmol/L compared with that in the control group (both P>0.05), but there were significant differences in the other experiment groups (all P<0.05). 
 Conclusion: JAK inhibitor AG490 can inhibit proliferation and promote apoptosis of the retinoblastoma HXO-RB44 cells through down-regulation of JAK2/STAT3 signaling pathway.

Objective: To evaluate the high-risk histopathologic features of retinoblastoma eyes enucleated as primary treatment. Methods: Retrospective case series study. An analysis was carried out on the high-risk histopathologic features of 482 primarily enucleated retinoblastoma eyes in Beijing Tongren Eye Center during December 2011 and December 2016. The high-risk histopathologic features included post lamina cribrosa invasion, massive choroidal invasion, anterior segment invasion and extraocular invasion. According to the neovascularization of iris (NVI) in the tissue specimen, the patients were divided into the NVI positive group and negative group. The rates of high-risk histopathologic features in the NVI positive group and negative group were analyzed using the chi-square test. Results: Among 482 children with retinoblastoma, 280 (58.1%) were males and 202 (41.9%) were females. The median age at enucleation surgery was 24 months (range, 1-84 months). Of the 482 primarily enucleated eyes, 184 (38.2%) were classified as high-risk histopathologic features, including 148 (30.7%) with post lamina cribrosa invasion, 15 (3.1%) with optic nerve transection invasion, 66 (13.7%) with massive choroidal invasion, 35 (7.3%) with anterior segment invasion, and 5 (1.0%) with extraocular invasion. NVI was detected in 257 (53.3%) cases and there was a significant relation between high-risk histopathologic features and NVI. The rate of high-risk histopathologic features was significantly higher in the NVI positive group (123 cases, 47.9%) than the NVI negative group (61 cases, 27.1%) (χ(2)=21.883, P= 0.001). The rate of post lamina cribrosa invasion was significantly higher in the NVI positive group (110 cases, 42.8%) than the NVI negative group (38 cases, 16.9%) (χ(2)=37.860, P= 0.001). Conclusions: In this large sample of patients, about 1/3 of retinoblastoma eyes has high-risk histopathologic features. The NVI positive retinoblastoma is more likely to have high-risk histopathologic features or post lamina cribrosa invasion than the NVI negative retinoblastoma. (Chin J Ophthalmol, 2018, 54: 782-786) .

In recent years, the treatment concept of retinoblastoma has changed from focusing on saving children's life to preserving the eyeballs and useful visual function. As a promising way to retain the eyeball, the application of vitrectomy in patients with retinoblastoma has long been debated. In view of the special anatomical structure of eyes and the biological characteristics of retinoblastoma cells, retinoblastoma is always prone to recurrence and metastasis after vitrectomy. Therefore, vitrectomy could hardly be a routine treatment and it should be used in retinoblastoma patients cautiously. (Chin J Ophthalmol, 2018, 54: 649-651) .

Retinoblastoma (RB) is the most common intraocular malignancy in childhood. It may seriously affect vision, and even threaten the life. The early diagnosis rate of RB in China remains low, and the majority of patients are at late phase with high rates of enucleation and mortality. The International Intraocular Retinoblastoma Classification and TNM staging system are guidances for therapeutic choices and bases for prognosis evaluation. Based on the sequential multi-method treatment modality, chemotherapy combined with local therapy is the mainstream in dealing with RB, which may maximize the results of eye saving and even vision retaining. New therapeutic techniques including supra-selective ophthalmic artery interventional chemotherapy and intravitreal chemotherapy can further improve the efficacy of treatment, especially the eye salvage rate. The overall level of RB treatment should be improved by promoting the international staging, new therapeutic techniques, and the sequential multiple modality treatment. (Chin J Ophthalmol, 2017, 53: 561 - 565) .

Intravenous chemotherapy and intra-arterial chemotherapy (IAC) both are the first-line treatment for retinoblastoma (RB) in clinical. There is a controversy on if intra-arterial chemotherapy can substitute the intravenous chemotherapy due to its high eye salvage rate in retinoblastoma therapy. The advantages and disadvantages of these two therapies were retrospectively reviewed here to suggest an individualized and comprehensive regimen for getting the proximal results for retinoblastoma afflicted children. (Chin J Ophthalmol, 2017, 53: 566 - 569) .

Objectives: To analyze RB1 gene mutation in retinoblastoma (RB) patients using gene capture technology. Methods: Experimental research. The clinical data of 17 RB patients were collected at Department of Ophthalmology, Peking University People's Hospital from June 2010 to Jun 2014. Peripheral blood samples of seventeen RB patients and their parents were collected and genomic DNA were extracted. DNA library from RB patients was mixed with designed gene capture probe of RB1 exons and its flanking sequences. The data were analyzed using bioinformatics software. To avoid the false positive, the abnormal sites were verified using the Sanger sequencing method. Results: Totally, there were 17 RB patients, including 12 males and 5 females, from 0.5 to 23 years old, average ages were (3.2±5.2) years old. Both eyes were involved in 6 patients. The other 11 cases were only one eye was attacked. Four RB patients were found to have germline mutations, among whom 2 had bilateral tumors and 2 had unilateral tumors. 2 novel missense mutations were identified, including 15(th) exon c.1408A>T (p. Ile470Phe) and c.1960G>C (p. Val654Leu) at 19(th) exon. No RB1 mutation was identified in any of their parents. We also identified 2 mutations reported previously. One is c.1030C>T termination mutation at 10(th) exon in a bilateral RB patients and his father, who was diagnosed with unilateral RB. The other is c.371-372delTA frame shift mutation at 3(rd) exon. No mutation was found in their parents. Conclusions: Two novel germline RB1 mutations were found using gene capture technology, which enriched RB1 mutations library. (Chin J Ophthalmol, 2017, 53: 455-459) .

Objective: To summarize the clinical features, treatment and prognosis of retinoblastoma (RB) patients in distant metastasis stage. Methods: Retrospective case series study. Clinical data of 24 cases (30 eyes) with diagnosis of distant metastasis stage RB were collected in pediatrics department of Beijing Tongren Hospital affiliated with Capital Medical University, from September 2005 to December 2013. In these cases, including 11 male cases and 13 female cases, the treatment age ranged from 7 months to 9 years with the median of 26 months. There were 18 unilateral cases (12 cases of right eye and 6 cases of left eye) and 6 bilateral cases. All cases were treated with surgery, chemotherapy and radiotherapy. The adverse reaction, blood routine and biochemical routine were monitored dynamically and regularly during treatment. The clinical manifestations and auxiliary examination were combined to evaluate the therapeutic effect. Determining the prognosis with death and survival, the survival curve and the median survival time were calculated by the product limit method. Results: The results of pathologic examination of 13 cases showed 11 cases of optic nerve involvement, 6 cases of optic nerve and choroid involvement, 1 case of post laminar involvement, 1 case of cornea and conjunctiva involvement. The most common metastatic site was central nerve system (CNS) (23 cases), followed by orbital involvement (10 cases). Other metastatic sites included bone (9 cases), bone marrow (1 case), pleura (1 case) and lymph node (1 case). 23 patients with CNS metastasis underwent systemic chemotherapy and intrathecal injection. The toxicity and side effects of radiotherapy and chemotherapy were evaluated and classified as grade I-II. Those adverse effects were tolerated by all cases. Followed up to December 31, 2015, all patients with CNS metastasis died and the median survival time was 6 months. Only 1 patient without CNS metastasis disease-free survived for 64 months. Conclusions: The most common metastasis site of RB was CNS. The overall prognosis of RB with distant metastasis was poor, in spite of given combined treatment mainly including surgery, chemotherapy and radiotherapy. But the prognosis of RB patients without CNS metastasis in distant metastasis stage was better than with CNS metastasis. (Chin J Ophthalmol, 2017, 53: 121-126) .

Purpose: To study the expression of matrix metalloproteinase-2(MMP-2), matrix metalloproteinase-9(MMP-9) and vascular endothelial growth factor(VEGF) in retinoblastoma (RB) and its relationship with the differentiation and optic nerve infiltration of RB., Methods: Immunohistochemical technique was used to detect the protein expression of MMP-2, MMP-9 and VEGF in 40 RB cases., Results: The MMP-2, MMP-9 protein and VEGF protein expression were detected as positive in 52.5%, 57.5% and 72.5% of 40 RB cases respectively. The expression of MMP-2, MMP-9 and VEGF was significantly higher in the undifferentiated pattern than in the rosetted pattern(p<0.05); and the expression of MMP-2, MMP-9 and VEGF was significantly higher in tumors with optic nerve invasion than in those without optic nerve invasion(p<0.05). In RB, the expression of MMP-2, MMP-9 had a positive correlation with the expression of VEGF., Conclusions: In RB, high expression of MMP-2, MMP-9 and VEGF is related to the invasion and metastasis of RB. The expression of MMP-2, MMP-9 is positively correlated with the expression of VEGF. The expression of MMP-2, MMP-9 and VEGF may act as a biological marker for the invasion and metastasis of RB. The method may help screen and identify the cases with high risk for metastasis and to predict the prognosis of patients.

Purpose: To investigate the expression of matrix metalloproteinase-1 (MMP-1) and vascular endothelial growth factor(VEGF) in the retinoblastoma and to identify their relationships with clinicopathology characteristics in order to determine their effects on the angiogenesis., Methods: Thirty-one patients with histologically proved retinoblastoma were enrolled to examine the expression of MMP-1and VEGF by Immunohistochemistry technique. The relationship between expression and the staging of tumor was analyzed., Results: The MMP-1 and VEGF were highly expressed in retinoblastoma, and MMP-1, VEGF mainly expressed in tumor cells. MMP-1and VEGF was positively correlated with tumor stage(p<0.05). The expression MMP-1 was positively correlated with VEGF level (R=0.787, p<0.00)., Conclusions: The MMP-1 and VEGF are highly expressed in retinoblastoma cells. The expression is related to clinical invasion and metastasis. MMP-1 expression is positively related to VEGF level.MMP-1and VEGF may play an important role in angiogenesis and tumor progression in retinoblastoma.

Objective: To develop a novel immunotherapy model of retinoblastoma (RB) in human PBL non-obese diabetic-severe combined immunodeficient (NOD-SCID) mice, and to investigate its biological features., Methods: Twenty NOD-SCID mice were randomly divided into 4 groups: group A, subcutaneous injection of phosphate buffered saline; group B, intraperitoneal injection of human peripheral blood lymphocyte (hu-PBL) for immune reconstruction; group C, subcutaneous injection of SO-RB50 cells and group D, intraperitoneal injection of hu-PBL with subcutaneous injection of SO-RB50 cells. The physical status of NOD-SCID mice and the tumor growth were observed. Human IgG in mouse serum was assayed by ELISA. Human T lymphocytes in murine spleen and peripheral blood were evaluated by flow cytometry. Histopathological examination and immunohistochemical studies were also performed., Results: In groups C and D, the latent period of tumor growth was 12 - 19 days, and the taken rates of RB were 100%. Histological and immunohistochemical results showed that the tumor cells of the two groups retained the characteristics of human low differentiation RB. More interesting, some human lymphocytes stained by human LCA presented in transplanted tumors and murine spleens. Human IgG and T lymphocytes were detected in humanized groups (groups B and D)., Conclusions: The RB model in the human PBL NOD-SCID mice has been successfully established, which simulates the biological behavior of human spontaneous RB. This model may be useful for the studying of immune regression and immunotherapy of human RB.