Apigenin Induced Apoptosis by Downregulating Sulfiredoxin Expression in Cutaneous Squamous Cell Carcinoma.

Cutaneous squamous cell carcinoma (cSCC) is the second carcinoma in nonmelanoma skin cancer (NMSC). Sulfiredoxin (Srx) is an antioxidant protein with a role in maintaining redox homeostasis. And Srx has an oncogenic role in skin tumorigenesis. In the current study, we found that apigenin, as a natural flavonoid, downregulated the expression of Srx protein in cSCC cell lines. Apigenin also inhibited the ability of cell proliferation and migration and induced apoptosis in cSCC cell lines. Our results also showed that apigenin induced apoptosis via the activation of the mitogen-activated protein kinase (MAPK) signaling pathway, as well as downregulated Srx expression in cSCC cell lines. Importantly, the effect of downregulation Srx by apigenin has been rescued with the inhibitor of the MAPK signaling pathway intervention. And induced apoptosis by apigenin was partially attenuated by the addition of MAPK inhibitor, Binimetinib. Our research revealed that apigenin induced apoptosis by downregulation of Srx expression through regulating the MAPK signaling pathway in cSCC cells, thus providing evidence of its applicability as a potentially effective therapeutic agent for cSCC treatment.

Effects of matrine on the proliferation and apoptosis of vincristine-resistant retinoblastoma cells.

Matrine is an active component of Leguminosae plants and is thought to exhibit anti-tumor effects. However, the effects of matrine on drug-resistant cancer have not been fully elucidated. The present study aimed to investigate the effects of matrine on vincristine (VCR)-resistant retinoblastoma (RB) cells and to assess the underlying mechanisms governing this effect. The drug-resistant cell line SO-Rb50/VCR was established by incubation with VCR at increasing concentrations. The effects of matrine on SO-Rb50 and SO-RB50/VCR cell growth and proliferation were evaluated using light microscopy and Cell-Counting Kit-8 assay. In addition, the effects of matrine on cell apoptosis, proliferation and cell cycle staging together with its potential underlying mechanisms were investigated. Matrine inhibited the proliferation of SO-Rb50 and SO-RB50/VCR cells in a concentration-dependent manner (0.2-1.1 mg/ml). However, matrine at the half-maximal inhibitory concentration (IC50) appeared to trigger apoptosis of these cells and had a tendency to arrest the cell cycle at the G0/G1 phase. Matrine treatment also promoted the expression of Bax and reduced the expression of Bcl-2 and cyclin D1 compared with the control. However, matrine was not able to increase the sensitivity of cells to VCR. The results of the present study suggested that matrine has the potential to promote the apoptosis of SO-Rb50/VCR cells and arrest cell cycling, indicating a possible benefit of matrine for the treatment of drug-resistant RB.

Detection of Circulating Tumor-Related Materials by Aptamer Capturing and Endogenous Enzyme-Signal Amplification.

Circulating tumor-related materials (CTRMs) shed from original or metastatic tumors, carry a lot of tumor information and are considered as important markers for cancer diagnosis and metastasis prognosis. Herein, we report a colorimetric detection strategy for CTRMs based on aptamer-based magnetic isolation and endogenous alkaline phosphatase (AP)-signal amplification. This strategy exhibited high sensitivity and selectivity toward the CTRMs that express AP heterodimers (the target of aptamer, a potential tumor marker). For clinical samples, this CTRM assay significantly discriminated colorectal cancer patients (n = 50) from healthy individuals (n = 39, p < 0.0001). The receiver operating characteristic (ROC) analysis indicated the sensitivity and specificity reached 92% and 82%, respectively, at the optimal cutoff point, the area under the curve of ROC reached 0.93, suggesting great potential for colorectal cancer diagnosis and therapeutic monitoring. Compared with CTC assays, this strategy is simple and has the potential for point-of-care testing.

Epstein-Barr virus-encoded latent membrane protein 1 promotes extracellular vesicle secretion through syndecan-2 and synaptotagmin-like-4 in nasopharyngeal carcinoma cells.

Increasing evidence indicates that extracellular vesicles (EVs) play an important role in cancer cell-to-cell communication. The Epstein-Barr virus (EBV)-encoded latent membrane protein 1 (LMP1), which is closely associated with nasopharyngeal carcinoma (NPC) pathogenesis, can trigger multiple cell signaling pathways that affect cell progression. Several reports have shown that LMP1 promotes EV secretion, and LMP1 trafficking by EVs can enhances cancer progression and metastasis. However, the molecular mechanism by which LMP1 promotes EV secretion is not well understood. In the present study, we found that LMP1 promotes EV secretion by upregulated syndecan-2 (SDC2) and synaptotagmin-like-4 (SYTL4) through nuclear factor (NF)-κB signaling in NPC cells. Further study indicated that SDC2 interacted with syntenin, which promoted the formation of the EVs, and SYTL4 is associated with the release of EVs. Moreover, we found that stimulation of EV secretion by LMP1 can enhance the proliferation and invasion ability of recipient NPC cells and tumor growth in vivo. In summary, we found a new mechanism by which LMP1 upregulates SDC2 and SYTL4 through NF-κB signaling to promote EV secretion, and further enhance cancer progression of NPC.

Correction: Lentivirus-Mediated Knockdown of Astrocyte Elevated Gene-1 Inhibits Growth and Induces Apoptosis through MAPK Pathways in Human Retinoblastoma Cells.

[This corrects the article DOI: 10.1371/journal.pone.0148763.].

LMP1-positive extracellular vesicles promote radioresistance in nasopharyngeal carcinoma cells through P38 MAPK signaling.

Radioresistance has been one of the impediments to effective nasopharyngeal carcinoma (NPC) therapy in clinical settings. Epstein-Barr virus (EBV) encoded latent membrane protein 1 (LMP1) is expressed in NPC and has potent effects on radioresistance. It has been detected in extracellular vesicles (EVs) or exosomes and shown to promote tumor proliferation and invasive potential. However, whether LMP1-positive EVs can confer radioresistance to cancer cells and the mechanism used to promote radioresistance need to be elucidated. In this study, the data showed that EVs derived from LMP1-positive NPC cells could induce recipient NPC cell proliferation and invasion and suppress apoptosis, especially promoting radioresistance. In addition, LMP1 could increase the secretion of LMP1-positive EVs. Furthermore, transmitted LMP1 subsequently performed its oncogenic functions through activating P38 MAPK signaling in recipient cells, and inhibiting P38 activity could efficaciously restore the sensitivity of NPC cells to ionizing radiation (IR). Finally, we found that LMP1-positive EVs could promote tumor growth and P38 inhibition eliminates this promoting effect in vivo, and EV formation is associated with a poor prognosis in NPC patients. These results showed that a few cells expressing LMP1 could enhance the radioresistance of NPC cells through potentially impacting the infected host and also modulating the tumor microenvironment.

EBV­LMP1 is involved in vasculogenic mimicry formation via VEGFA/VEGFR1 signaling in nasopharyngeal carcinoma.

The Epstein-Barr virus latent membrane protein 1 (EBV­LMP1) is an oncoviral protein that plays an important role in oncogenic transformation in EBV­associated nasopharyngeal carcinoma (NPC). Our previous studies demonstrated that LMP1 increased VEGFA expression and upregulated angiogenesis in NPC. Vasculogenic mimicry (VM) is a mechanism by which tumor cells can obtain nutrients to survive, and VM has been observed in numerous types of tumors. However, the occurrence and significance of VM in NPC and the relationship between LMP1 and VM have not yet been evaluated. In the present study, we observed that it was almost impossible for LMP1-negative NPC cells to form tubular structures, whereas LMP1-positive NPC cells were able to form tubular structures. Moreover, VEGFA was found to be involved in VM formation in LMP1-positive NPC cells. Knockdown of LMP1 or VEGFR1 distinctly disrupted tubular structures, whereas inhibition of VEGFR2 did not affect the process, indicating that VEGFR1 not VEGFR2 signaling was involved in LMP1-mediated VM formation. Furthermore, the data of immunohistochemistry (IHC) and CD34/PAS double staining in a tumor tissue array showed that LMP1 was positively correlated with VEGFR1 and VM. Meanwhile, after analyzing the clinicopathological features, we found that VM formation was associated with a poor prognosis in NPC patients. These results suggest that VM formation is increased by EBV­LMP1 via VEGF/VEGFR1 signaling and provide additional information to clarify the role of EBV­LMP1 in NPC pathophysiology.

Association between choroidal thickness and anterior chamber segment in eyes with narrow or open-angle.

AIM: To investigate the relationship between choroidal thickness and anterior chamber segment in subjects with eyes with narrow or open-angle. METHODS: The subfoveal choroidal thickness was measured with enhanced depth-imaging optical coherence tomography and anterior chamber parameters were measured with ultrasound biomicroscopy in one eye of 23 subjects with open-angle eyes and 38 subjects with narrow-angle eyes. The mean age was 59.52±7.04y for narrow-angle subjects and 60.76±7.23y for open-angle subjects (P=0.514). Multivariate linear regression analysis was performed to assess the association between choroidal thickness and narrow-angle parameters. RESULTS: There were no differences in subfoveal choroidal thickness between open- and narrow-angle subjects (P=0.231). Anterior chamber parameters, including central anterior chamber depth, trabecular iris angle, iris thickness 500 µm from the scleral spur (IT500), and ciliary body thickness at 1 mm and 2 mm from the scleral spur (CBT1, CBT2) showed significant differences between the two groups (P<0.05). Subfoveal choroidal thickness showed negative correlation (ß=-0.496, P=0.016) only with anterior chamber depth in the open-angle group and with age (ß=-0.442, P=0.003) and IT500 (ß=-0.399, P=0.008) in the narrow-angle group. However, subfoveal choroidal thickness was not correlated with trabecular iris angle, anterior chamber depth, ciliary body thickness, or central corneal thickness in the narrow-angle group. CONCLUSION: Choroidal thickness does not differ in the two groups and has not correlated with anterior chamber parameters in narrow-angle subjects, suggesting a lack of relationship between choroidal thickness and primary angle-closure glaucoma.

Scleral and choroidal volume in relation to axial length in infants with retinoblastoma versus adults with malignant melanomas or end-stage glaucoma.

PURPOSE: To measure scleral and choroidal volume in eyes of Chinese, and to assess associations with age and axial length. METHODS: We histomorphometrically examined globes from infants and adults which had been enucleated due to retinoblastoma, uveal melanoma, or absolute painful glaucoma. Thickness of sclera and choroid were measured, and volumes were calculated. RESULTS: The study included 225 globes (mean axial length: 24.6 ± 4.2 mm; range:17.0-35.7 mm; mean age: 30.4 ± 22.6 years; range: 1-83 years). Mean computed scleral volume was 648 ± 136 mm(3). Scleral volume in children aged <5 years significantly increased with longer axial length (P = 0.001; correlation coefficient r: 0.42) and older age (P = 0.003; r: 0.39) in univariate analysis. In multivariate analysis within the group of children aged ≤2 years, larger scleral volume increased with longer axial length (P = 0.04; standardized correlation coefficient beta: 0.32; correlation coefficient B: 21.6; 95 % confidence interval (CI): 0.52, 42.7) and showed a statistically non-significant tendency to increase with older age (P = 0.06;b eta: 0.30; B: 56.9; 95% CI: -1.5,115). In individuals aged ≥ 5 years, scleral volume was not significantly associated with axial length (P = 0.75) or age (P = 0.13). Mean choroidal volume as measured and calculated in 95 individuals (age: 16-81 years) was 44.1 ± 14.1 mm(3), and was not significantly associated with age (P = 0.47; r: -0.08) or axial length (P = 0.83; r: -0.02). CONCLUSIONS: This study on children eyes with retinoblastoma and adult eyes with malignant melanomas or end-stage glaucoma suggests that primary eye growth up to an age of 2 years is associated with an increase in scleral volume. After the age of 2 years, scleral volume and choroidal volume remain unchanged, leading to scleral and choroidal thinning with longer axial length, in particular at the posterior pole.

Lentivirus-Mediated Knockdown of Astrocyte Elevated Gene-1 Inhibits Growth and Induces Apoptosis through MAPK Pathways in Human Retinoblastoma Cells.

PURPOSE: To explore expression and function of astrocyte elevated gene-1 (AEG-1) in human retinoblastoma (RB). METHODS: The expression of AEG-1 in histological sections of human RBs and in RB cell lines was examined using immunohistochemical staining and RT-PCR and Western blotting respectively. We knocked down AEG-1 gene levels by AEG-1-siRNA lentivirus transfection of human RB cell lines SO-RB50 and Y79, and using an MTT assay, we assessed the role of AEG-1 on RB cell proliferation. The biological significance of lentivirus transfection induced AEG-1 down-regulation was examined by assessing the apoptosis rate in the transfected RB cells by Annexin V-APC staining and flow cytometry. We additionally measured the expression of Bcl-2, Bax, cleaved-caspase-3 and caspase-3, and the phosphorylation and non-phosphorylation alternation of MAPKs. RESULTS: AEG-1 expression was detected to be strongly positive in the histological slides of 35 out of 54 (65%) patients with RB. AEG-1 expression increased significantly (P<0.05) with tumor stage. In the RB cell lines SO-RB50, Y79 and WERI-RB1 as compared with retinal pigment epithelium cells, expression of AEG-1 mRNA and AEG-1 protein was significantly higher. In AEG-1-siRNA lentivirus transfected cell cultures as compared with negative control lentivirus transfected cell cultures, levels of AEG-1 mRNA and of AEG-1 protein (P<0.05) and cell growth rates (P<0.01) were significantly lower, and apoptosis rate (P<0.001), Bax/Bcl-2 ratio and cleaved-caspase-3 protein level were significantly increased. The P-ERK/ERK ratio was significantly decreased in the AEG-1-siRNA lentivirus transfected cell lines. CONCLUSIONS: Expression of AEG-1 was associated with RB, in histological slides of patients and in cell culture experiments. Lentivirus transfection induced knockdown of AEG-1 had a tumor suppressive effect, potentially by tumor cell apoptosis induction through inhibition of ERK.

SCLERAL AND CHOROIDAL THICKNESS IN SECONDARY HIGH AXIAL MYOPIA.

PURPOSE: To assess differences in scleral and choroidal thickness between eyes with secondary high axial myopia caused by congenital glaucoma, eyes with primary high axial myopia, and nonhighly myopic eyes. METHODS: The study consisted of 301 Chinese individuals with a mean age of 23.9 ± 22.6 years and mean axial length of 24.8 ± 4.2 mm. It included the "secondary highly myopic group" (SHMG) because of congenital glaucoma (n = 20 eyes; axial length >26.0 mm), the "primary highly myopic group" (PHMG) (n = 73; axial length >26.0 mm), and the remaining nonhighly myopic group (NHMG). RESULTS: The secondary highly myopic group versus the primary highly myopic group had significantly thinner sclera in the pars plana region (343 ± 71 µm versus 398 ± 83 µm; P = 0.006), whereas scleral thickness in other regions did not differ significantly between both highly myopic groups and was significantly thinner in both highly myopic groups than in the NHMG. Mean total scleral volume did not differ significantly (P > 0.20) between any group (SHMG: 659 ± 106 µm; PHMG: 667 ± 128 µm; NHMG: 626 ± 135 µm). Choroidal thickness was significantly thinner in both highly myopic groups than in the NHMG, with no significant differences between both highly myopic groups. Choroidal volume did not differ significantly (P > 0.40) between any of the groups (SHMG: 43 ± 12 µm; PHMG: 43 ± 13 µm; NHMG: 46 ± 17 µm). CONCLUSION: In secondary high axial myopia, the sclera gets thinner anterior and posterior to the equator; whereas in primary high axial myopia, scleral thinning is predominantly found posterior to the equator. Because volume of sclera and choroid did not differ between any group, scleral and choroidal thinning in myopia may be due to a rearrangement of tissue and not due to the new formation of tissue.

The receptor proteins: pivotal roles in selective autophagy.

Autophagy is a highly regulated and multistep biological process whereby cells under metabolic, proteotoxic, or other stresses remove dysfunctional organelles and/or misfolded/polyubiquitinated proteins by shuttling them via specialized structures called autophagosomes to the lysosome for degradation. Although autophagy is generally considered to be a non-selective process, accumulating evidence suggests that it can also selectively degrade specific target cargoes. These selective targets include proteins, mitochondria, and even invading bacteria. The discovery and characterization of autophagic adapters, such as p62/Sequestosome 1 (SQSTM1) and Neighbor of BRCA1 gene 1 (NBR1), have provided mechanistic insights into selective autophagy. These receptors are all able to act as cargo receptors for the degradation of ubiquitinated substrates. This review mainly summarizes the most up-to-date findings regarding the key receptor proteins that play important roles in regulating selective autophagy.

Retinoblastoma in Chinese Children Aged Five to Fourteen Years.

OBJECTIVE: To assess the characteristics of retinoblastomas enucleated from Chinese children aged 5-14 years. METHODS: This retrospective hospital-based study included all eyes with retinoblastomas consecutively enucleated in the Beijing Tongren Hospital between August 2003 and July 2013. RESULTS: Out of 1,205 patients, 47 (3.9%) were 5 years or older. All tumors in this age group occurred unilaterally, the patients had a negative family history, and the tumors were detected at an age of 6.9 ± 1.8 years (range: 5-14). The main clinical features at the time of examining the as yet untreated children aged 5-7 years (n = 30) or >7-14 years (n = 10) were leukocoria, strabismus, pseudohypopyon, hypertension, vitreous seeds ('snowballs'), and calcifications. In 12 patients (26%), the retinoblastoma had not initially been diagnosed as a tumor. Histopathology revealed tumor invasion into nonretinal tissue in 19 eyes (40%). Therapy included enucleation only (n = 22; 47%), adjuvant systemic chemotherapy (n = 24; 51%), and additional orbital exenteration (n = 1). After a mean follow-up of 3.0 ± 2.1 years (range: 0.2-9.8), which was done for 40 children, none of these children showed a tumor recurrence. CONCLUSIONS: Of the children undergoing enucleation for retinoblastoma in Beijing, 3.9% were aged 5 years or more. As in Western countries, the tumor occurrence was unilateral, their family history was negative, and the survival rate was relatively high in these children. In 1 out of 4 children, the tumor had initially been misdiagnosed due to a masquerade syndrome. Retinoblastoma should be considered in the differential diagnosis of any unclear intraocular situation in children.

Scleral Thickness in Chinese Eyes.

PURPOSE: We measured scleral thickness in eyes of Chinese, and assessed interregional differences and associations with age and axial length. METHODS: Using light microscopy, we histomorphometrically measured scleral thickness at various locations in eyeballs from Chinese patients that had been enucleated due to retinoblastoma, uveal melanoma, or absolute painful glaucoma. RESULTS: The study included 281 globes from patients with a mean age of 24.8 ± 23.1 years (range, 1-83 years) and mean axial length of 24.3 ± 3.9 mm (range, 17.0-35.7 mm). In multivariate analysis in children aged ≤2 years, thicker posterior scleral thickness was marginally significantly associated with older age (P = 0.07; standardized correlation coefficient ß, 0.21; correlation coefficient B, 62.2; 95% confidence interval [CI], -4.3,128.8) after adjusting for shorter axial length (P = 0.01). In participants aged ≥5 years, larger posterior scleral thickness was significantly associated only with shorter axial length (P < 0.001; ß, -0.49; B, -24.1; 95% CI, -30.6,-17.6), but not with age (P = 0.93). The ratio of posterior scleral thickness to scleral thickness in the pars plana region decreased significantly with increasing axial length. The correlation coefficients were higher for the ratio of scleral thickness posterior pole/pars plana region than for posterior pole/ora serrata, posterior pole/equator, or posterior pole/midpoint posterior pole to equator. Scleral thickness measurements were not significantly (all P > 0.10) associated with adult glaucoma. CONCLUSIONS: Scleral thickness increased up to an age of 2 years, while afterwards scleral thickness was independent of age and decreased with longer axial length. Differences in the associations between the regional scleral thickness ratios and axial length suggested a scleral thinning taking place in axially elongated eyes predominantly in the posterior globe segment.

[Retinoblastoma spontaneous regression: clinical and histopathologic analysis].

OBJECTIVE: To analyze the clinicopathologic characteristics of retinoblastoma spontaneous regression. METHODS: Retrospective case series study. The clinic and pathologic data of 52 cases of retinoblastoma spontaneous regression were analyzed retrospectively. All the cases came from 579 cases of retinoblastoma (without pre-enucleation chemotherapy) archived in Ophthalmic Pathology Laboratory of Beijing Tongren Eye Centre from Jan. 2007 to Dec. 2013. The analyzed factors included tumor activity, tumor invasion, and tissue changes of uveal and lens. RESULTS: In this study, the patients' average age was (14.0 ± 8.7) months. There were 33 (63.5%) male and 19 (36.5%) female patients. The rate of retinoblastoma spontaneous regression was 9.0% (52/579), of which the rate of complete regression was 1.4% (8/579). The most common symptom was leucocoria coupled with red eye 55.8% (29/52). The typical clinicopathologic characteristics were as follows: (1) all or most of all tumor cells suffered coagulation necrosis and only tumor cell remains were left; (2)most eyeballs showed atrophiabulbi or phthisis bulbi; (3) severe atrophy occurred in uvea, especially in iris and ciliary body combined with pigment cells disintegrating; (4) cataract formed coupled with lens swelling, displacement, or wrapped by fibrous connective tissue; (5) in a small number of cases, there were alive tumor cells in the retrolaminar optic nerve or choroid in spite of most tumor cells had suffered regression, indicating the patients had histopathologic high risk factors and had the dispositions of tumor diffusion and metastasis. CONCLUSIONS: Some untypical clinical manifestation such as leukocoria coupled with red eye or atrophia bulbi should be paid close attention. The possibility of retinoblastoma spontaneous regression should be considered. Meanwhile, some cases of retinoblastoma spontaneous regression with histopathologic high risk factors should be given post-enucleation systemic adjuvant chemotherapy combined with close observations and follow-ups.

Oncogenic GNAQ and GNA11 mutations in uveal melanoma in Chinese.

PURPOSE: To examine whether GNAQ and GNA11 somatic mutations previously identified in uveal melanomas of Caucasians are associated with uveal melanomas in Chinese patients. METHODS: Uveal melanomas treated by primary enucleation in Chinese patients underwent a mutation analysis of GNAQ and GNA11 with sequencing of exon 5 and exon 4. RESULTS: The study included 50 patients with uveal melanoma and with a mean age of 47.6±13.0 years. During the follow-up of at least 3 years, 20 (40%) patients developed extraocular metastases. The frequencies of GNAQ and GNA11 somatic mutations in uveal melanoma were 18% (9/50) and 20% (10/50), respectively. The mutations occurred exclusively in codon 209 of exon 5. No mutations were detected in exon 4. Mutations affecting codon 209 in GNAQ were c.626A>C(Q209P) (78%) and c.626A>T(Q209L) (22%). Mutations affecting codon 209 in GNA11 were exclusively c.626A>T(Q209L) (100%). In none of the tumors, mutations of BRAF and NRAS were detected. GNAQ/11 mutations were marginally (P = 0.045) associated with optic disc involvement. In Kaplan-Meier analysis, metastasis-free survival was not significantly (P = 0.94) associated with GNAQ/11 mutations. CONCLUSIONS: Mutations of GNAQ and GNA11 can be found in Chinese patients as in Caucasian patients with uveal melanoma, with a higher frequency reported for Caucasian patients.

Bovine serum albumin nanoparticles as controlled release carrier for local drug delivery to the inner ear.

Nanoparticles have attracted increasing attention for local drug delivery to the inner ear recently. Bovine serum albumin (BSA) nanoparticles were prepared by desolvation method followed by glutaraldehyde fixation or heat denaturation. The nanoparticles were spherical in shape with an average diameter of 492 nm. The heat-denatured nanoparticles had good cytocompatibility. The nanoparticles could adhere on and penetrate through the round window membrane of guinea pigs. The nanoparticles were analyzed as drug carriers to investigate the loading capacity and release behaviors. Rhodamine B was used as a model drug in this paper. Rhodamine B-loaded nanoparticles showed a controlled release profile and could be deposited on the osseous spiral lamina. We considered that the bovine serum albumin nanoparticles may have potential applications in the field of local drug delivery in the treatment of inner ear disorders.