Luteolin inhibits the proliferation, adhesion, migration and invasion of choroidal melanoma cells in vitro.

Choroidal melanoma is a devastating disease that causes visual loss and a high mortality rate due to metastasis. Luteolin, a potential anticancer compound, is widely found in natural plants. The aim of this study was to evaluate the antiproliferative, antiadhesive, antimigratory and anti-invasive effects of luteolin on choroidal melanoma cells in vitro and to explore its potential mechanism. Cell counting kit-8 (CCK-8) assays, 5-ethynyl-2'-deoxyuridine (EdU) assays, Cell adhesion, migration, and invasion assays were performed to examine the inhibitory effects of luteolin on cell cell viability, proliferation, adhesion, migration and invasion capacities, respectively. Considering the correlation between Matrix metalloenzymes and tumor metastasis, Enzyme-linked immunosorbent assays (ELISAs) were used to assess matrix metalloproteases MMP-2 and MMP-9 secretion. Western blotting was performed to detect p-PI3K P85, Akt, and p-Akt protein expression. The cytoskeletal proteins vimentin were observed with cell immunofluorescence staining. Luteolin can inhibit OCM-1 cell proliferation, migration, invasion and adhesion and C918 cell proliferation, migration, and invasion through the PI3K/Akt signaling pathway. Therefore, Luteolin may have potential as a therapeutic medication for Choroidal melanoma.

The role of Cdk5-mediated Drp1 phosphorylation in Aß1-42 induced mitochondrial fission and neuronal apoptosis.

Alzheimer's disease, one of the most common neurodegenerative diseases, is pathologically characterized by Amyloid beta containing plaques and neurofibrillary tangles. Amyloid beta (Aß) induces neuronal apoptosis through the intracellular Ca2+ increase, subsequent hyperactivation of cyclin-dependent kinase 5 (Cdk5) and mitochondrial abnormality. Recently, Cdk5 was identified as an upstream regulator of mitochondrial fission during neuronal apoptosis, but the underlying mechanism remains unclear. Here, in vitro phosphorylation assays showed that Cdk5 could phosphorylate the recombinant Drp1 at Serine 579. Aß1-42 stimulation increased the phosphorylation level of Drp1 at Serine 579 in mouse cortical neurons. Cdk5 inhibitor roscovitine and knockdown of Cdk5 by a lentiviral vector expressing shRNA targeting Cdk5 (Lenti-Cdk5-shRNA) efficiently prevented Aß1-42 induced Drp1 phosphorylation in neurons. In addition, Aß1-42 stimulation induced markedly mitochondrial fission in neurons. Roscovitine, Lenti-Cdk5-shRNA and expression of phospho-defect mutatant GFP-Drp1-S579A in neurons attenuated Aß1-42 induced mitochondrial fission, whereas expression of phospho-mimetic mutant GFP-Drp1-S579D alone resulted in mitochondiral fission similar to Aß1-42 stimulation. Moreover, Roscovitine and Lenti-Cdk5-shRNA suppressed the cleavage of caspase-3 and protected neurons against Aß1-42 induced neuronal apoptosis.Thus, our data indicate that Drp1 is a direct target of Cdk5, and Cdk5-mediated phosphorylation of Drp1 at Serine 579 regulates Aß1-42 induced mitochondrial fission and neuronal toxicity.

Effect of navigation endoscopy combined with three-dimensional printing technology in the treatment of orbital blowout fractures.

AIM: To explore the combined application of surgical navigation nasal endoscopy (NNE) and three-dimensional printing technology (3DPT) for the adjunctive treatment of orbital blowout fractures (OBF). METHODS: Retrospective analysis was conducted on the data of patients with OBF who underwent surgical treatment at the Affiliated Eye Hospital of Nanchang University between July 2012 and November 2022. The control group consisted of patients who received traditional surgical treatment (n=43), while the new surgical group (n=52) consisted of patients who received NNE with 3DPT. The difference in therapeutic effects between the two groups was evaluated by comparing the duration of the operation, best corrected visual acuity (BCVA), enophthalmos difference, recovery rate of eye movement disorder, recovery rate of diplopia, and incidence of postoperative complications. RESULTS: The study included 95 cases (95 eyes), with 63 men and 32 women. The patients' age ranged from 5 to 67y (35.21±15.75y). The new surgical group and the control group exhibited no statistically significant differences in the duration of the operation, BCVA and enophthalmos difference. The recovery rates of diplopia in the new surgical group were significantly higher than those in the control group at 1mo [OR=0.03, 95%CI (0.01-0.15), P<0.0000] and 3mo [OR=0.11, 95%CI (0.03-0.36), P<0.0000] post-operation. Additionally, the recovery rates of eye movement disorders at 1 and 3mo after surgery were OR=0.08, 95%CI (0.03-0.24), P<0.0000; and OR=0.01, 95%CI (0.00-0.18), P<0.0000. The incidence of postoperative complications was lower in the new surgical group compared to the control group [OR=4.86, 95%CI (0.95-24.78), P<0.05]. CONCLUSION: The combination of NNE and 3DPT can shorten the recovery time of diplopia and eye movement disorder in patients with OBF.

Biomechanical analysis of an absorbable material for treating fractures of the inferior orbital wall.

AIM: To investigate the biomechanical properties and practical application of absorbable materials in orbital fracture repair. METHODS: The three-dimensional (3D) model of orbital blowout fractures was reconstructed using Mimics21.0 software. The repair guide plate model for inferior orbital wall fracture was designed using 3-matic13.0 and Geomagic wrap 21.0 software. The finite element model of orbital blowout fracture and absorbable repair plate was established using 3-matic13.0 and ANSYS Workbench 21.0 software. The mechanical response of absorbable plates, with thicknesses of 0.6 and 1.2 mm, was modeled after their placement in the orbit. Two patients with inferior orbital wall fractures volunteered to receive single-layer and double-layer absorbable plates combined with 3D printing technology to facilitate surgical treatment of orbital wall fractures. RESULTS: The finite element models of orbital blowout fracture and absorbable plate were successfully established. Finite element analysis (FEA) showed that when the Young's modulus of the absorbable plate decreases to 3.15 MPa, the repair material with a thickness of 0.6 mm was influenced by the gravitational forces of the orbital contents, resulting in a maximum total deformation of approximately 3.3 mm. Conversely, when the absorbable plate was 1.2 mm thick, the overall maximum total deformation was around 0.4 mm. The half-year follow-up results of the clinical cases confirmed that the absorbable plate with a thickness of 1.2 mm had smaller maximum total deformation and better clinical efficacy. CONCLUSION: The biomechanical analysis observations in this study are largely consistent with the clinical situation. The use of double-layer absorbable plates in conjunction with 3D printing technology is recommended to support surgical treatment of infraorbital wall blowout fractures.

Tumor subtypes and signature model construction based on chromatin regulators for better prediction of prognosis in uveal melanoma.

Background: Uveal Melanoma (UM) is the most prevalent primary intraocular malignancy in adults. This study assessed the importance of chromatin regulators (CRs) in UM and developed a model to predict UM prognosis. Methods: Gene expression data and clinical information for UM were obtained from public databases. Samples were typed according to the gene expression of CRs associated with UM prognosis. The prognostic key genes were further screened by the protein interaction network, and the risk model was to predict UM prognosis using the least absolute shrinkage and selection operator (LASSO) regression analysis and performed a test of the risk mode. In addition, we performed gene set variation analysis, tumor microenvironment, and tumor immune analysis between subtypes and risk groups to explore the mechanisms influencing the development of UM. Results: We constructed a signature model consisting of three CRs (RUVBL1, SIRT3, and SMARCD3), which was shown to be accurate, and valid for predicting prognostic outcomes in UM. Higher immune cell infiltration in poor prognostic subtypes and risk groups. The Tumor immune analysis and Tumor Immune Dysfunction and Exclusion (TIDE) score provided a basis for clinical immunotherapy in UM. Conclusion: The risk model has prognostic value for UM survival and provides new insights into the treatment of UM.

Luteolin Suppresses Three Angiogenesis Modes and Cell Interaction in Uveal Melanoma in Vitro.

PURPOSE: Uveal melanoma is a high-vascularized tumor that lacks effective systemic therapies. Most anti-angiogenesis drug therapies only target endothelial cell-dependent angiogenesis but not vasculogenic mimicry (VM), which supplies blood to tumors independent of endothelial cells. Thus, we aimed to explore the inhibitory effects of luteolin on proliferation, migration, invasiveness, angiogenesis, and VM activity of uveal melanoma. We further explored the signaling pathway underlying the mechanism of action of luteolin. METHODS: Monocultures of uveal melanoma C918 cells, human umbilical vein endothelial cells (HUVECs), and co-cultures of these two cell lines were established. Angiogenesis of HUVECs, VM formation of C918 cells, and the mosaic vessels formed by both cell types were observed under an inverted microscope. Cell counting kit-8, 5-ethynyl-2'-deoxyuridine (EdU), wound scratch, Transwell cell migration, and invasion assays were performed. VEGF levels were detected by ELISA. Western blotting was used to detect the expression of PI3K, p-PI3K P85, Akt, and p-Akt Ser473 proteins. RESULTS: Luteolin inhibited all three modes of angiogenesis observed in uveal melanoma in vitro. Luteolin effectively inhibited the proliferation, migration, and invasion of C918 cells and proliferation and migration of HUVECs. Furthermore, luteolin could inhibit the interaction between the endothelial cells and C918 cells. VEGF secretion in C918 cells and HUVECs treated with luteolin was inhibited. Luteolin decreased the levels of phosphorylated Akt kinase. CONCLUSION: We demonstrated the anti-angiogenic effects of luteolin, including against the VM type, in addition to suppressing tumor cell proliferation and migration in vitro. Furthermore, luteolin likely exerts its inhibitory effects via the phosphatidylinositol 3-kinase (PI3K)/protein kinase B (AKT) signaling pathway. Luteolin might be an effective therapeutic candidate for treating highly vascularized uveal melanoma tumors.

Biological role of postoperative low level laser therapy in preventing hydroxyapatite orbital implantation exposure: A case report.

Conjunctival sac stenosis is the contraction of the conjunctival sac as a result of trauma or disease. The aim of the present study was to observe the clinical effects of low-level laser therapy (LLLT) combined with hydroxyapatite (HA) orbital implantation as a treatment strategy for conjunctival sac stenosis. A total of 10 patients with conjunctival sac stenosis were treated with scleral graft transplantation in conjunction with HA implantation and postoperative LLLT. In addition, a rabbit model was used to investigate the biological mechanism underlying the effects of LLLT with the aim of preventing and treating orbital implantation exposure. The right eyeball was removed, orbital implantation performed and LLLT applied to experimental groups. 99mTc-Methyl diphosphonate scanning methods were performed at different timepoints to compare the average radioactivity count of the region of interest between surgical (right) and control (left) eyes (R/L). Histopathological examination was performed 8 weeks post-surgery, followed by analysis of fiber vascularization. Following LLLT, moderate conjunctival wounds were completely healed within 2 weeks and severe stenosis wounds healed within 3 weeks. Following prosthesis implantation in the rabbit model, a significantly elevated R/L ratio was observed after 4 weeks, whereas no significant difference was observed compared with the control group at 6 and 8 weeks postoperatively. Histopathological examination revealed that all implants were fibrotic. Overall, the present study demonstrated that LLLT promoted the survival of conjunctival grafts, stimulated conjunctival incision healing and promoted early vascularization of HA implants. Clinical trial registration no: ChiCTR-DDT-12002660 (www.chictr.org/cn/).

Effect of luteolin on apoptosis and vascular endothelial growth factor in human choroidal melanoma cells.

AIM: To investigate the effects of luteolin on apoptosis, the cell cycle, and the expression and secretion of vascular endothelial growth factor (VEGF) in human choroidal melanoma cells (C918 and OCM-1). METHODS: C918 and OCM-1 cells cultured in vitro were treated with various concentrations of luteolin (0, 5, 10, 15 µmol/L). Cell growth was observed with an inverted microscope, and cell cycle arrest was detected by propidium iodide (PI) staining using flow cytometry. Apoptosis was detected by Hoechst33342 staining, and apoptosis rate was determined by Annexin V-FITC/PI experiments using flow cytometry. The expression of apoptosis-related proteins Bcl-2, Bax and VEGF was analyzed using Western blots. The levels of VEGF secreted by the cells into the supernatant was analyzed using ELISA. RESULTS: After treating with 5 to 15 µmol/L luteolin for 48h, the fusion degree of C918 and OCM-1 cells decreased, and more floating apoptotic cells appeared. Luteolin treatment increased the G0-G1 phase ratio of the C918 and OCM-1 cells, blocked cell cycle progression, and increased the apoptosis rate of the C918 and OCM-1 cells. Western blot showed that luteolin decreased the expression of Bcl-2 and VEGF in the C918 and OCM-1 cells and increased the expression of Bax protein. The ELISA results showed that 10 to 15 µmol/L luteolin decreased the cell secretion of VEGF. CONCLUSION: Luteolin may induce apoptosis by regulating the levels of apoptosis-related proteins in C918 and OCM-1 cells. Luteolin can induce cell cycle arrest, decrease the expression of VEGF.

Three-year clinical observation of the outcomes of transepithelial and epithelial-off accelerated corneal collagen crosslinking treatment for different types of progressive keratoconus.

In the present study, the clinical and long-term effects of accelerated transepithelial corneal collagen crosslinking (ATE-CXL) and accelerated epithelial-off corneal collagen crosslinking (A-CXL) for the treatment of different types of progressive keratoconus were compared. A total of 70 patients, including 96 eyes with advanced keratoconus, were enrolled in the study. ATE-CXL or A-CXL was performed on one or two eyes of each subject according to corneal thickness, keratoconus type and surgical approach. Patients were divided into the following four groups: Group A, ATE-CXL for central keratoconus; group B, A-CXL for central keratoconus; group C, ATE-CXL for peripheral keratoconus; and group D, A-CXL for peripheral keratoconus. Uncorrected distant visual acuity (UDVA), best-corrected distant (BD)VA and corneal astigmatism (CA) were evaluated in all patients by routine ophthalmology pre-operatively and 3 years post-operatively. Topographical features, including maximum corneal curvature (Kmax), thinnest corneal thickness (TCT), anterior corneal elevation (ACE) and corneal endothelial cell density (ECD) were also compared across groups. The results suggested that pre- and post-operative UDVA, BDVA, Kmax, CA and ACE values differed in all four groups (P<0.05), whereas no differences were observed between pre- and post-operative TCT and ECD (P>0.05). Concordant results were obtained between groups A and C and groups B and D. ATE-CXL achieved better control of central keratoconus UDVA, Kmax and CA as compared with A-CXL. The difference between pre- and post-operative UDVA, Kmax and CA as compared with A-CXL was highly correlated with the change in intraocular pressure and treatment effectiveness. There was a statistically significant improvement in BDVA with ATE-CXL for treatment of central keratoconus compared with that after A-CXL treatment (P=0.032). There were statistically significant improvements in BDVA (P=0.047), CA (P=0.045) and ACE (P=0.012) with A-CXL treatment of peripheral keratoconus when compared with ATE-CXL treatment. Central, and to a lesser extent, peripheral, keratoconus may be effectively controlled by either approach, with disease stabilization 3 years later. ATE-CXL is suggested to be the most suitable treatment for keratoconus of <400 µm with a corneal thickness of >400 µm; however, A-CXL yields superior long-term outcomes.

Analysis of the effect of repair materials for orbital blowout fracture on complications.

AIM: To investigate the effect of repair materials for orbital blowout fractures on the occurrence of postoperative complications. METHODS: The clinical data and follow-up data of 54 subjects with orbital blowout fractures were retrospectively analyzed. The study was divided into three groups according to the used repair materials: titanium mesh (16 cases), Medpor (12 cases), and Medpor titanium mesh (26 cases). All test data were analyzed using the SPSS version 23.0 statistical software. The mean age and duration of disease between the groups were compared through one-way analysis of variance. The Chi-square (χ 2) test was used to compare the number of males and females, different fracture types, and different surgical approaches among groups. The χ 2 test was used to compare the frequencies for complications in each group. RESULTS: The baseline characteristics of age and gender in each group were matched (F=1.763, P=0.172; χ 2=0.026, P=0.987). In addition, there was no difference in the type of fracture and surgical approach (χ 2=0.460, P=0.977; χ 2=0.691, P=0.952), or the incidence of complications (χ 2=0.081, P=0.960) between the three groups. CONCLUSION: Although there is no difference in effect of various repair materials on the incidence of complications, the effect of repair materials on postoperative complications of orbital blowout fractures should not be ignored.

Acetylcholinesterase function in apoptotic retina pigment epithelial cells induced by H2O2.

AIM: To investigate the acetylcholinesterase (AChE) expression involved in retina pigment epithelial (RPE) apoptosis induced by higher concentrations H2O2. METHODS: The human retinal pigment epithelium cell line ARPE-19 was from ATCC (Rockville, MD). Cultured ARPE-19 cells were treated with H2O2 at 0, 250, 500, 1 000, 2 000µmol/L and cell viability was measured with MTT assay. AChE expression and DNA fragments were analyzed by immunocytochemistry, TUNEL and PARP-1 Western blotting. RESULTS: Immunofluorescence detected AChE exist in the normal human retinal tissue. When H2O2 >500µmol/L, AChE expression showed an increase after 2h, and this concentration was selected for the present study. RPE cell was induced with 1 000µmol/L H2O2 for 2h, compared to the control group, cell activity decline detected by MTT, AChE and PARP-1 protein expression was significantly increased detected by Western blotting. AChE immunofluorescence staining was positive in RPE cell after H2O2 incubate 2h. In addition, pretreatment with 100µmol/L epigallocatechin gallate (EGCG), cell viability increased from 31.20%±3.90% to 70.23%±12.96%. CONCLUSION: AChE is weakly expressed in normal human RPE cells. Stimulation with H2O2 caused the stable increase of AChE expression in RPE cells, which may indicate that AChE may be an important role in AMD.

[Semiconductor low level laser irradiation for exposure of hydroxyapatite orbital implants].

OBJECTIVE: To evaluate the efficacy of semiconductor low level laser irradiation for the treatment of postoperative exposure of hydroxyapatite orbital implants. METHODS: 22 cases with postoperative exposure of hydroxyapatite orbital implants were divided into three groups according to the size of implants exposure. The exposure wound in the 3 groups was irradated with semiconductor low level laser 5 min per day for 5-15 days. The follow-up period ranged from 2 to 24 months. RESULTS: In the group with less then 3 mm of exposure, the wound healed in 1 week after 5-10 days irradiation; in the group with implant exposure of 4-7 mm, the would healed in 1-2 weeks after 10-15 days irradiation; in the group with implant exposure of 8-10 mm, the would healed in 2-3 weeks after 10-15 days irradiation. Compared with the treatments of drugs and/or surgical repair, which was used for another 20 cases of exposure of hydroxyapatite orbital implants, semiconductor low level laser increased healing rate obviously in the groups with implant exposure of 4-7 mm and 8-10 mm (P = 0.019, 0.018). CONCLUSION: Semiconductor low level laser has better effects than drugs and/or surgical repair for exposure of hydroxyapatite orbital implants.