Objective To investigate inhibited effects of melatonin (MLT) on proliferative activity of retinoblastoma cell line HXORB44 and its related mechanism. Methods HXO-RB44 cells were treated by MLT of different concentration (10-10, 10-9, 10-8, 10-7 mmol/L. Cell counting and tetrazolium dyereduction assay (MTT) were used to determine the effect of MLT on the survival and proliferation of HXO-RB44 cells. Apoptotic nuclei were further analyzed by HoechstPI fluorescence staining. Flow cytometry was used to measure the fluorescent intensity of ROS, cell cycle distribution and apoptosis. Results 10 -6 mmol/L (or exceed) of MLT could inhibit the proliferation of HXO-RB44cells in vitro while 10-7 mmol/L (or below) of MLT couldn't. With the increase of MLT concentration from 10-10 mmol/L to 10-7 mmol/L, HXO-RB44 cells gradually increased the expression of ROS. Hoechst staining showed that 4, 8, 12 and 24 hours after the incubation with MLT, the nuclear pyknosis and nuclear fragmentation increased in HXORB44 cells. The extent of apoptosis was proportional to the concentrations of MLT. Flow cytometry revealed that with the increasing of MLT concentration, G0/G1 and G2/M phase cells increased, S phase cells decreased. The apoptotic rate was also increased. Conclusion 10 -6 M of MLT could inhibit the proliferation of HXO-RB44 cells. This effect may relate to the increased ROS expression, cell cycle arrest at G0/G1 phase and apoptosis of HXO-RB44 cells.
OBJECTIVE：Observing the clinical and pathological features of Coats disease. METHODS:Reviewing the clinical data and pathologic slides duly confirmed by pathology of 19 cases of Coats disease,which belonging to our college's Laboratory of Ophthalmologic Pathology from 1959 to 1994. RESULTS: 14 males,5 females,aged 1-18 years. More boys were affected than girls in the age group under 10 and that difference between both sexes became gradually less as they grew older. The main pathologic changes were the vascular dilatation and congestion of the outer layer of the retina,the uneven thickness of the vascular walls and the proliferation of the connective tissue. Retinal protuberance was seen in most of the advanced cases.with bleeding and vascular changes on its surfaces. The main pathologic changes were the detachment of retina and the appearance of many foam cells and crystals of cholesterol in the subretnal fluid,and calcification and ossification of the outer layer of the retina were found in some cases. CONCLUSION :Cytological examination of the subretinal fluid might be the liable method in differentiating between the Coats disease and retinoblatstoma. （Chin J Ocul Fundus Dis,1996,12: 157-159）
A review of 426 patients with retinoblastoma(RB)who presented to Eye Hospital of Zhongshan Ophthalmic Center from 1966 to 1991 showed that 81(19.01%)patients were more than or equal to 5 years of age(median age 6.95 years)at the time of initial diagnosis.All patients are unilateral except 4 cases who are bilateral.The main presentations included leukocoria,exophthalmos,congertion and pain of the eye,drcreased vision,strabismus,oculare hypertension,vitreous opacity,hypopyon and hyphema,etc.Some atypical presentations in these older patients with RB were discussed and some problems that should be noticed in the diagnosis and management were presented.discussed and some problems that should be noticed in the diagnosis and management were presented. （Chin J Ocul Fundus Dis,1993,9:205-206）
Objective To investigate the effect of exogenous Rb gene on the cell cycle of vitreous retinoblastoma (RB) transplantation tumor in nude mouse. Methods Based on establishing vitreous RB transplantation tumor in nude mouse,constructing retrovirus vector of Rb gene PBabe-Rb and transfecing it into the RB transplantation model by liposome Dosper,the change of cell cycle of the RB transplantation tumor by flow cytometry(FCM)was analysed． Results FCM showed that the cells of G1phase of the treated eyes were obviously more than the control eyes with the value of DNA index(DI)and S phase fraction(SPF) decreased by the Rb gene expression. Conclusion The exogenous PBabe-Rb gene can partially suppress the progress of the cell cycle of RB transplantation tumor in vivo. (Chin J Ocul Fundus Dis，2000，16：1-70)
PURPOSE:To investigate the relationship between the proliferative activity of refinoblastoma (RB)cell and the RB differentiation degree and the infiltration capability. METHOD:The proliferating cell nuclear antigen (PCNA)expression in RB tissues of 48 cases was analysed by using LSAB immunohistochemical method. RESULTS :The mean PCNA labelling index(LI)in differentiated RB tissues of 12 cases was markedly lower than that in non-differentiated of 36 cases(P＜0.05). The mean PCNA LI in RB tissues of the optic nerve infiltrated group(22 cases)was significantly higher than that of the optic nerve non-infiltrated group(26 cases)(P＜0.05). The results indicate that the PCNA LI is significantly related with the differentiation degree of RB and the infiltration capability. CONCLUSION :The determination of PCNA LI is of significance for evaluating the histologic characteristics and biological behavior of RB.
Objective To investigate the effect of doxycycline on the proliferation and vasculogenic mimicry in retinoblastoma (RB) cell line in vitro. Methods RB cell line were tested for their ability to form perfusable tubular networks in 3D culture with doxycycline in the concentrations ranging from 5 to 20 mg/L, and CoCl2 was used as chemical hypoxia-inducing reagent to mimic tumor hypoxic microenvironment. The effect of doxycycline on proliferation were detected by MTT assay in vitro, and the effect on tube formation of RB cells were detected by tube-like structure formation assay and PAS staining. The mRNA levels of MMP2 and MMP9 at different hypoxic culture and different doxycycline concentrations were detected by semiquantitative reverse transcription polymerase chain reaction (RT-PCR). Results The micrograph showed that RB cells linked each other to form cavity and network tructure in 3D culture. the number of tubules in doxycycline group were significantly lower than which in the control group in the concentrations ranging from 5 to 20 mg/L (Plt;0.001).OD of doxycycline group was significantly lower than which in the control group (t=15.320，Plt;0.01) , The proliferation of RB cells had a negative correlation with the concentration of doxycycline (r =-0924, Plt;001). The levels of MMP2 and MMP9 mRNA of RB cells under hypoxia were significantly higher than which in the control group (t=16.469，Plt;0.01). As the concentration of doxycycline increased, the expression of MMP-2 and MMP-9 decreased. The result of double staining also showed that VM, formed by CD34negative and PASpositive tumor cells, existed in 12 simples of retinoblastoma. Conclusion RB cells have the capacity of selfmetamorphosing and vasculorizing in 3D culture. Doxycycline can inhibit their proliferation and vasculogenic mimicry formation in vitro by downregulating the expression of MMP-2 and MMP-9 .
Objective To explore the expression of survivin gene in retinoblastoma (RB) and its relationship with the stages and histodifferentiation degree of RB and the expression of p53、bcl-2 proteins. Methods Expression of survivin, p53 and bcl-2 proteins in 38 RB conventional paraffin samples were detected with survivin, p53 and bcl-2 monoclonal antibodies respectively by immunohistochemical assay. The expression of survivin of normal retina in 6 control samples was observed. Results In 38 cases of RB, positive expression of survivin was found in 20 (52.6%); while none of the 6 normal retinal tissue expressed survivin, which had significant difference between the two group (P＜0.05). The positive expression of survivin did not correlate with sex of patient, disease stages and histological type (P＞0.05). In 38 RB cases, positive expression of p53 was in 25 with the rate of 65.8%, and of bcl-2 in 18 with the rate of 47.4%. The positive-expressed rates were much higher in positive-expressed p53 and bcl-2 group than those in the negative-expressed p53 and bcl-2 group(P＜0.05). Conclusion The increase of the expression of survivin implies that it may take part in the occurrence and development of RB; the interaction among survivin, p53 and bcl-2 may participate in the access and the course of RB. (Chin J Ocul Fundus Dis，2004，20：215-217)
Objective To observe the expression of sperm protein (SP) 32/OY-TES-1 in retinoblastoma (RB).Methods Thirty paraffin specimens of pathologically confirmed RB eyeballs were investigated in this study. The SP32/OY-TES-1 mRNA expression of 15 RB tissues, 12 non-tumor retinal tissues and 22 normal eye tissues were detected by reverse transcription polymerase chain reaction (RT-PCR). The SP32/OY-TES-1 protein expression of 30 RB tissues and 24 normal retinal tissues were examined by immunohistochemistry. The relationship between SP32/OY-TES-1 mRNA, protein expression and age, gender, tumor size, tumor differentiation, clinical stage of RB children were analyzed. Results The expression rate of SP32/OY-TES-1 mRNA in RB tissues, non-tumor retinal tissues and normal eye tissues were 86.7%, 16.7% and 36.4% respectively. Compared the expression rate of SP32/OY-TES-1 mRNA in different gender (chi;2=0.744)，age（chi;2=1.178），clinical stage(chi;2=1.188)，tumor size (chi;2=0.216)，tumor differentiation(chi;2=1.885)，the differences were not statistically significant (P＞0.05). The expression rate of SP32/OY-TES-1 protein in RB tissues was 73.3% and no expression in normal retinal tissues. Compared the expression rate of SP32/OY-TE-1 protein in different gender (chi;2=1.000)，agewith le;two years and ＞two years（chi;2=0.403），tumor size (chi;2=2.274)，tumor differentiation(chi;2=0.138), the differences were not statistically significant (P＞0.05); but in clinical stage (chi;2=6.193)，with or without optic nerve infiltration (chi;2=4.535), the differences were statistically significant(P＜0.05). Conclusions SP32/OY-TES-1 is highly expressed in RB. The expression rate of SP32/OY-TES-1 is related to optic nerve infiltration and clinical stage of RB.
ObjectiveTo observe the clinical characteristics and survival rate of the patients with extraocular retinoblastoma (RB). MethodsThis is a retrospective case analysis. From November 2003 to May 2015, 38 eyes of 31 patients with RB in the extra-ocular stage from 213 RB patients were enrolled in this study. There were 18 males and 13 females. Bilateral lesions were observed in 7 patients and unilateral lesions were observed in 24 patients.19 patients were diagnosed at less than 2 years old, 10 patients at 2 to 5 years old, and 2 patients at age over 5 years old. First visit time was less than 1 month in 12 patients, from 1 to 3 months in 15 patients, over 3 months to 6 months in 4 patients. Medical history and family history were record at the first visit. All patients underwent orbital CT, MRI, double color Doppler imaging and wide angle digital retinal imaging system. CT and (or) MRI examination detected tumor extraocular invasion. Histopathological examination showed that there were tumor cells invasion of the scleral, optic nerve root and optic nerve. Chemotherapy was done after surgery. In the extra-ocular stage, 3 to 6 rounds of intensive chemotherapy combined with orbital radiotherapy were done. The average follow-up period was (25.5±4.5) months after treatment. The cumulative survival rate was observed after 6 months, 1 and 5 years after treatment, and the relationship between the initial age, time, sex, single eye, tumor and survival time of the patients was analyzed. ResultsThe extraocular RB accounted 14.55% of all RB patients in this study. There is no family history of RB, no special history. There were 15 patients with leukocoria and yellow-white reflection in the pupil; 5 patients with lacrimation, swelling, photophobia and exophthalmos; 11 patients with strabismus. The cumulative survival rate at 6 months, 1, 5 years after treatment was (78.0±9.0)%, (62.0±11.0)%, (57.0±11.0)% respectively. The average survival time was (53.9±7.8) months; the cumulative survival rate was (59.3±11.3)%. When the age of first visit was less than 1 month, 1-3 months, 3-6 months, the median survival time was 78, 15 and 18 months respectively, the cumulative survival rate was 100.0%, (40.0±21.9)% and (25.0±21.7)%, respectively. The survival time of the newly diagnosed patients at 1 month was more than at 1 to 6 months, and the difference was statistically significant (t=9.20, P < 0.05). Conclusions14.55% of all RB patients was extraocular RB in this study. One of the most common clinical manifestations is leukocoria at the first visit. The cumulative survival rate of extraocular RB is lower, while the survival rate of patients with the age of first visit time was less than 1 month is higher.
Objective To study the relation between retinoblastoma (Rb) gene expression and biological characteristics of gallbladder carcinoma. Methods The expression of Rb protein in tissues of 41 cases of gallbladder carcinoma, 7 cases of gallbladder papilloma, and 14 cases of cholecystitis were detected by immunohistochemical staining of SP with polyclonal antibody. Results The rate of positive staining was 58.7% in gallbladder carcinoma which was significantly lower than that in cholecystitis and gallbladder papilloma (100%). There was a significant difference of Rb protein expression among the low, moderate and high differentiations and so was between S5, S4, S1, S2 and S3 stage (Ｐ＜0.05). Conclusion It sugests that expression or non-expression of Rb gene is closely related to the malignant potential invasiveness and prognosis of gallbladder carcinoma.