Objective To observe the clinical features of congenital hypertrophy of retinal pigment epithelium （CHRPE）. Methods The clinical data of 13 CHRPE patients including visual acuity, slit-lamp microscope examination, indirect ophthalmoscope examination and fundus fluorescein angiography (FFA) were retrospectively analyzed. The patients, 9 males and 4 females, with the mean age of 27.8 years. Results All patients were unilateral, without systemic diseases and no subjective symptoms in majority. Only 30.77% of initial diagnosis was correct, other diagnosis include choroidal nevi, old chorioretinopathy or no diagnosis. The round or oval black lesion was found in ocular fundus of all patients, 7.69% was located on the optic disk, 46.15% was located on the inferior temporal retina, 30.77% was located on the superior temporal retina, 15.39％ was located on the inferior nasal retina. 92.31％ was pigmented CHRPE and 7.69％ was non-pigmented CHRPE. FFA showed blocked fluorescence and transmitted fluorescence in the lesion, few eyes were found dilated capillary vessel and fluorescent leakage on the late stage of FFA, most eyes had normal retinal vessels. Conclusion The isolated CHRPE is round or oval black lesion in ocular fundus which lack of subjective symptoms, mostly located on the peripheral retina; the FFA characteristics showed blocked fluorescence and transmitted fluorescence, and CHRPE often misdiagnosed as other disease, it should be combine the ocular fundus manifestation with the FFA to diagnose properly.
Objective To evaluate the effect of Schwann cell (SC) on the proliferation of marrow mesenchymal stem cells (MSCs) and provide evidence for application of SC in construction of the tissue engineered vessels.Methods SC and MSCs were harvested from SD rats(weight 40 g). SC were verified immunohstochemically by the S-100 staining, and MSCs were verified by CD 44, CD 105, CD 34 and CD 45. The 3rd passages of both the cells were cocultured in the Transwell system and were amounted by the 3H-TDR integration technique at 1, 3, 5 and 7 days,respectively. The results were expressed by the CPM(counts per minute, CPM) values. However, MSCs on both the layers were served as the controls. The Westernblot was performed to assess the expression of the vascular endothelial growth factor (VEGF), its receptor Flk-1, and the associated receptor neuropilin 1(NRP-1) in SC, the trial cells, and the controls. Results SC had a spindle shape in the flasks, and more than 90% of SC had a positive reaction for the S-100 staining.MSCs expressed CD44 and CD105, and had a negativesignal in CD 34 and CD 45. The CPM values of MSCs in the trial groups were 2 411.00±270.84，3 016.17±241.57，6 570.83±2 848.27 and 6 375.8±1 431.28at 1, 3, 5 and 7 days, respectively. They were significantly higher in their values than the control group (2 142.17±531.63，2 603.33±389.64，2 707.50±328.55，2 389.00±908.01), especially at 5 days （P＜0.05）. The Western blot indicated that VEGF was expressedobviously in both the SC group and the cocultured MSCs grou,p and was less visible in the control cells. The expressions of Flk-1 and NRP-1 inthe cocultured MSCs were much ber than in the controls. Conclusion SC can significantly promote the proliferation of MSCs when they are cocultured. The peak time of the proliferation effect appeared at 5 days. This effect may be triggered by the up-regulation of VEGF in MSCs, which also leads to the upregulation of Flk-1 and NRP-1 .
Objective To investigate the effect of berbamine (BBM) on the proliferation and apoptosis of retinoblastoma (RB) HXO-RB44 cells and its possible mechanism in vitro.Methods RB cells in logarithmic growth phase were divided into BBM treated group and control group. RB cells in BBM treated group were cultured with different concentrations of BBM (2,4,8,16 and 32 mg/L) for 24,48 and 72 hours, respectively. The proliferation was assayed by methyl Thiazolyl tetrazolium (MTT). RB cells were cultured with different concentrations of BBM (4,8 and 16 mg/L) for 24 hours. The early apoptotic rates were detected by flow cytometry; the expression of bcl-2 and Bax were measured by enzyme-linked immunosorbent assay (ELISA) and the activity of Caspase-3 was detected by colorimetric assay.Results BBM could obviously inhibit the proliferation of RB cells in a time and dose dependent manner (24 hours: F=70.547,P＜0.01; 48 hours: F=603.438,P＜0.01; 72 hours: F=577.521,P＜0.01). The IC50 value at 24,48 and 72 hours were 25.26, 10.94 and 6.25 mg/L, respectively. Necrosis rates of control group and BBM treated group were (1.25plusmn;0.45)%, (4.10plusmn;2.95)%, (4.39plusmn;0.21)% and (10.54plusmn;4.38)% respectively; the difference between two groups was statistically significant (F=6.527,P＜0.05). Apoptotic and necrosis rates in advanced stage of control group and BBM treated group were (2.13plusmn;0.71)%, (5.45plusmn;2.31)%, (9.86plusmn;3.18)% and (11.10plusmn;1.70)%, respectively. The difference between two groups was statistically significant (F=10.845,P＜0.05). Early apoptotic rates of control group and BBM treated group were (0.51plusmn;0.26)%, (2.68plusmn;0.35)%, (5.97plusmn;0.50)% and (11.22plusmn;1.17)%, respectively. The difference between two groups was statistically significant (F=144.976,P＜0.01). In addition, BBM dose-dependently reduced bcl-2 level and increased Bax expression, causing the reduction of the bcl-2/Bax protein ratio as well as increased the Caspase-3 activity in RB cells remarkably (bcl-2: F=835.726,P＜0.01; bax: F=111.963, P＜0.01;Caspase-3:F=298.058,P＜0.01).Conclusions BBM can inhibit the proliferation and induce apoptosis or necrosis of RB cells in vitro, down regulating the expression of bcl-2, up regulating the expression of Bax. Along with increased Caspase-3 activity these may be the apoptotic mechanisms.
Objective To investigate the relationship of the expression between heat shock protein (HSP) 70 and 90, and Survivin and its effects on the proliferative activity in retinoblastoma (RB) cells. Methods Expression of Survivin, HSP70 and 90, and Ki-67 in conventional paraffin samples from 43 patients with RB and 6 healthy people was detected by streptavidin-biotin peroxidase (SP) immunohistochemical method. Ki67 labeling index was used to evaluate the proliferative activity in RB. Results In 43 cases of RB, positive expression of HSP70 and 90 and Survivin was found in 28 (65.12%), 37 (86.05%) and 27 (62.79%) cases, respectively. None of the 6 normal retinal tissue expressed HSP70, HSP90 or Survivin. Positive expression of Survivin was more frequent in positive expressions of HSP90 than that in negative expressions of HSP90 (P＜0.05). Ki67 labeling index was higher in positive expressions of HSP90 and positive expressions of Survivin than that in their negative expressions respectively (P＜0.05). Meanwhile, higher Ki67 labeling index was found in positive HSP90Survivin expressions than that in negative HSP90Survivin expressions and those cases where only HSP90 or Survivin was found (P＜0.05). Expression of HSP70 did not correlate with that of Survivin, nor had any significant effect on Ki67 labeling index (P＞0.05). Expression of HSPs and Survivin and Ki67 labeling index did not correlate with histological types (P＞0.05). Conclusion Expression of HSP90 correlates with that of Survivin in RB. Co-existence of Survivin and HSP90 probably plays an important role in the genesis of RB.
Objective To study the effects of adenosine 2A receptor activation on activation, proliferation, and toxicity of T lymphocytes stimulated by phytohemagglutinin (PHA) in vitro. Methods A model of activated T cells was established by stimulating the cells with PHA. Those T cells were treated with different concentrations of adenosine 2A receptors agonist (0.01 μmol/L, 0.1 μmol/L, 1 μmol/L, and 10 μmol/L CGS21680). The expressions of CD69, CD25 and proliferation of T cells were measured by fluorescent antibody stain and flow cytometry. ELISA method was used to detect IL-2 and INF-γ levels. Results All concentrations of CGS21680 significantly inhibited the expressions of CD25 and CD69 on PHA-stimulated T cells surface and proliferation of T cells （Plt;0.05， Plt;0.01）. IL-2 and INF-γ secreted by T cells were significantly suppressed, too （Plt;0.01）. Conclusion Activation of adenosine 2A receptor can effectively inhibit the activation, proliferation, and toxicity of T cells in vitro.
Objective To study the effects of L-arginine （L-Arg） on cell proliferation, inducible nitric oxide synthase (iNOS) expression and cell cycle in human colon carcinoma cell line LS174 through nitric oxide (NO) pathway. Methods LS174 cells were cultured in medium with L-Arg at different concentrations for different times. MTT method was employed to evaluate the level of the cell proliferation. The production of NO in culture supernatants of LS174 cell was detected with enzyme reduction of nitrate. The distribution of the cell cycle was detected with the flow cytometry (FCM). The expression level of iNOS in the cells was determined by Western blot and SP immunocytochemical staining method. Results The growth of LS174 was promoted by the L-Arg at low concentration (0.125 mmol/L) and inhibited at high concentrations (0.5， 2， 8 and 32 mmol/L). The level of NO was increased with the increasing concentration of L-Arg in culture medium. To compare with the control group, the ratio of cells at S phase was increased after 48 hours’ treatments with high concentrations (0.5, 2, 8 and 32 mmol/L) of L-Arg (P＜0.05, P＜0.01); while there was no obvious difference after treatments with low concentration (0.125 mmol/L) of L-Arg (Pgt;0.05). With the increase of the concentration of L-Arg, the expression of iNOS was increased as compared with control group. The higher the concentration of L-Arg was, the better the effect. Conclusion L-Arg can induce the expression of iNOS resulting in increase the production of nitric oxide (NO). Low concentration of L-Arg can promote the growth of LS174 cells, while high concentration ones can inhibit growth and proliferation. The high concentration of L-Arg could induce S phase arrestion in the cell cycle.
Objective To investigate the impacts of neoadjuvant chemotherapy on the expression of insulin-like growth factor-1 receptor (IGF-1R) and on operation procedure and the significance of prognosis. Methods The expression of IGF-1R in 40 patients with breast cancer before and after neoadjuvant chemotherapy was measured by immunohistochemistry. The diagnosis was proved by core biopsy. All the patients took the TAC chemotherapy regimen. Modified radical operation was performed after two chemotherapy cycles and the IGF-1R expression was measured again. The clinical effect of neoadjuvant chemotherapy was assessed according to WHO criterion by measuring the size of tumor by physical examination and B type ultrasound. Results After neoadjuvant chemotherapy the tumor size shrank in 29 patients, there was no CR (complete response) or PD (progressed disease) to be documented. IGF-1R expression could be downregulated in 25 patients. Conclusion Neoadjuvant chemotherapy can inhibit the tumor growth by downregulation of the expression of IGF-1R.
Objective To investigate the effect of ursolic acid on the proliferation and apoptosis of human osteosarcoma cell line U2-OS and analyze its mechanism. Methods Human osteosarcoma cell line U2-OS was divided into 4 groups, which was cultured with ursolic acid of 0, 10, 20, and 40 μmol/L, respectively. At 0, 24, 48, and 72 hours after being cultured, the cell proliferation ability was detected by cell counting kit 8 (CCK-8). At 48 hours, the effects of ursolic acid on cell cycle and apoptosis of U2-OS cells were measured by flow cytometry. Besides, the expressions of cyclin D1 and Caspase-3 were detected by real-time fluorescent quantitative PCR and Western blot. Results CCK-8 tests showed that the absorbance (A) value of each group was not significant at 0 and 24 hours (P>0.05); but the differences between groups were significant at 48 and 72 hours (P<0.05). Flow cytometry results showed that, with the ursolic acid concentration increasing, the G1 phase of U2-OS cells increased, the S phase and G2/M phase decreased, and cell apoptosis rate increased gradually. There were significant differences between groups (P<0.05). Compared with the 0 μmol/L group, the relative expressions of cyclin D1 mRNA and protein in 10, 20, and 40 μmol/L groups significantly decreased (P<0.05); whereas, there was no significant difference in relative expression of Caspase-3 mRNA between groups (P>0.05). However, with the ursolic acid concentration increasing, the relative expressions of pro-Caspase-3 protein decreased and the relative expressions of activated Caspase-3 increased; there were significant differences between groups (P<0.05). Conclusion Ursolic acid can effectively inhibit the proliferation of osteosarcoma cell line U2-OS, induce the down-regulation of cyclin D1 expression leading to G0/G1 phase arrest, increase the activation of Caspase-3 and promote cell apoptosis.
ObjectiveTo observe apoptosis and proliferation of choledochus wall epithelial cell and fibrocyte, to understand the effects of apoptosis and proliferation on choledochal cyst development.MethodsThirty two cases of cystic dilatation,35 cases of cylindrical dilatation，and 25 cases of cholangiectasis caused by choledocholith were collected. All specimens were offered by department of hepatobiliarypediatric surgery. The apoptosis related index (bcl2 and bax) and cell proliferation index (PCNA) were detected by the immunohistochemical technique; Apoptosis was detected by TUNEL method. ResultsThere was serious mucosal epithelial cell damage in cystic dilatation group. In cylindrical dilatation group there was a damage similar to that of the cystis dilatation group, but the damage was not serious. In control group there was little damage in the duct wall, but there was a low positive rate of apoptosis of 〔epithelium cell (2.74±1.00)% and fibroblast (2.95±0.87）%〕, and a low bcl2 and bax’s expression rate, and a high PCNA’s expression rate 〔epithelium cell (3.74±1.00)%, fibroblast (3.71±1.77)%〕. There was no obvious difference between cylindrical dilatation group and cystic dilatation group (Pgt;0.05): the PCNA’s expression rate was low 〔(0.99±0.51)% and (0.90±0.38)% respectively〕, the bax expression rate was high in remaining epithelial cell, and the positive rate of bax was apparently higher than that of bcl2 (P＜0.05), the positive rate of the apoptosis cell was high 〔(13.94±4.77)%, (7.51±3.46)%〕; the expression rate PCNA were high 〔(9.91±2.91)%,(9.70±3.18)%〕, and expression rate of bax’s was low in the fibre tissue, the positive rate of bcl2 was markedly higher than that of bax, and the positive rate of the apoptosis cell was low 〔(3.74±2.12)%,(4.46±2.41)%〕. There were no marked difference between the two groups (Pgt;0.05). The expression of bcl2 and bax had marked difference both in cylindrical dilatation group and cystic dilatation group and as compared to control group (P＜0.05). ConclusionApoptosis has certain promoting effect in the course of choledochal cyst formation.
【Abstract】ObjectiveTo investigate the growth effect of methylprednisolone （MP） on human hepatocellular carcinoma cell line HepG2.Methods Periodic distribution of cells and cellular apoptosis were detected by using cell culture,immunofluorescence staning of Annex Ⅴ and flow cytometric analysis in hepatocellular carcinoma cell.Results Compared with control group, methylprednisolone increased Ｇ0/Ｇ1 phase cell, decreased S phase cell on human hepatocellular carcinoma cell line HepG2 ,which had positive correlation with the time.The apoptosis rate and the necrosis rate of cells had the relation of dose-dependent with the concentration of MP, the cell membrane of early cellular apoptosis was stained green fluorescence. Conclusion Methylprednisolone can induce Ｇ0/Ｇ1 arrest , may play a proliferation-inhibition effect on the hepatocellular carcinoma cell line HepG2.