Brca1 Is Regulated by the Transcription Factor Gata3, and Its Silencing Promotes Neural Differentiation in Retinal Neurons.

Previous studies have indicated that Brca1 (Breast cancer suppressor gene 1) plays an important role in neural development and degenerative diseases. However, the bioactivity and regulatory mechanism of Brca1 expression in retinal neurocytes remain unclear. In the present study, our data indicated that Brca1 maintains the state of neuronal precursor cells. Brca1 silencing induces differentiation in 661W cells. Nestin, a marker of precursor cells, was significantly decreased in parallel with Brca1 silencing in 661W cells, whereas Map2 (Microtubule associated protein 2), a marker of differentiated neurons, was significantly increased. Neurite outgrowth was increased by ~4.0-fold in Brca1-silenced cells. Moreover, DNA affinity purification assays and ChIP assays demonstrated that Gata3 (GATA binding protein 3) regulates Brca1 transcription in 661W cells. Silencing or overexpressing Gata3 could significantly regulate the expression of Brca1 and affect its promoter inducibility. Furthermore, the expression of Gata3 generally occurred in parallel with that of Brca1 in developing mouse retinas. Both Gata3 and Brca1 are expressed in the neonatal mouse retina but are developmentally silenced with age. Exogenous Gata3 significantly inhibited neural activity by decreasing synaptophysin and neurite outgrowth. Thus, this study demonstrated that Brca1 is transcriptionally regulated by Gata3. Brca1/Gata3 silencing is involved in neuronal differentiation and maturation.

Alpha-1-antichymotrypsin as a novel biomarker for diagnosis, prognosis, and therapy prediction in human diseases.

The glycoprotein alpha-1-antichymotrypsin (AACT), a serine protease inhibitor, is mainly synthesized in the liver and then secreted into the blood and is involved in the acute phase response, inflammation, and proteolysis. The dysregulation of AACT and its glycosylation levels are associated with tumor progression and recurrence, and could be used as a biomarker for tumor monitoring. In this review, we summarized the expression level, glycosylation modification, and biological characteristics of AACT during inflammation, neurodegenerative or other elderly diseases, and tumorigenesis, as well as, focused on the biological roles of AACT in cancer. The aberrant expression of AACT in cancer might be due to genetic alterations and/or immune by bioinformatics analysis. Moreover, AACT may serve as a diagnostic or prognostic biomarker or therapeutic target in tumors. Furthermore, we found that the expression of AACT was associated with the overall survival of patients with human cancers. Decreased AACT expression was associated with poor survival in patients with liver cancer, increased AACT expression was associated with shorter survival in patients with pancreatic cancer, and decreased AACT expression was associated with shorter survival in patients with early lung cancer. The review confirmed the key roles of AACT in tumorigenesis, suggesting that the glycoprotein AACT may serve as a biomarker for tumor diagnosis and prognosis, and could be a potential therapeutic target for human diseases.

The link between diabetic retinal and renal microvasculopathy is associated with dyslipidemia and upregulated circulating level of cytokines.

Purpose: To investigate the mechanisms underlying the correlations between diabetic retinopathy (DR) and diabetic nephropathy (DKD) and examine whether circulating cytokines and dyslipidemia contribute to both DR and DKD in patients with 2 diabetes mellitus (T2DM). Methods: A total of 122 patients with T2DM were enrolled and categorized into the DM group (without no DR and DKD), DR group [non-proliferative DR (NPDR), and proliferative DR (PDR)] with no DKD), DR complicated with DKD groups (DR+DKD group). The biochemical profile, including fasting blood glucose (FBG), glycated hemoglobin (HbA1c), and lipid profile were estimated, and plasma inflammatory and angiogenic cytokines [monocyte chemoattractant protein-1 (MCP-1), interleukin (IL)-6, IL-8, vascular endothelial growth factor (VEGF)-A, C, D, and placental growth factor (PlGF)] were analyzed by protein microarrays. The atherogenic plasma index (API) was defined as low-density lipoprotein cholesterol (LDL-C)/high-density lipoprotein-cholesterol (HDL-C); atherogenic index (AI) was calculated as [(total cholesterol (TC) -HDL-C)/HDL-C], and atherogenic index of plasma (AIP) was defined as log (TG/HDL-C). Results: By multivariable disordered regression analysis, after controlling for duration of DM and hypertension, LDL-C (p = 0.019) and VEGF-D (p = 0.029) resulted as independent risk factors for DR. Albumin-to-creatinine ratio (uACR) (p = 0.003) was an independent risk factor for DR with DKD. In DR, NPDR, and PDR groups, grades of A1, A2, and A3 of albuminuria increased with the severity of DR. In A1, A2, and A3 grade groups, the severity of DR (DM, NPDR, and PDR) increased with higher albuminuria grades. Kendall's tau-b correlation coefficient analysis revealed that FBG (p = 0.019), circulating level of PlGF (p = 0.002), and VEGF-D (p = 0.008) were significantly positively correlated with the grades of uACR (p < 0.001), and uACR grades were significantly correlated with DR severity (p < 0.001). Conclusions: The occurrence and severity of DR are closely correlated with kidney dysfunction. Among the three kidney functional parameters, uACR resulted as the better indicator of DR severity and progression than glomerular filtration (eGFR) and serum creatinine (Scr). Impaired FBG was associated with microalbuminuria, emphasizing that well-controlled FBG is important for both DR and DKD. The link between diabetic retinal and renal microvasculopathy was associated with dyslipidemia and upregulated circulating level of angiogenic cytokines.

An In Vitro Model of Diabetic Retinal Vascular Endothelial Dysfunction and Neuroretinal Degeneration.

BACKGROUND: Diabetic retinopathy (DR) is a leading cause of blindness in working-age populations. Proper in vitro DR models are crucial for exploring pathophysiology and identifying novel therapeutic targets. This study establishes a rational in vitro diabetic retinal neuronal-endothelial dysfunction model and a comprehensive downstream validation system. METHODS: Human retinal vascular endothelial cells (HRMECs) and retinal ganglion cells (RGCs) were treated with different glucose concentrations with mannitol as matched osmotic controls. Cell proliferation and viability were evaluated by the Cell Counting Kit-8. Cell migration was measured using a transwell migration assay. Cell sprouting was assessed by a tube formation assay. The VEGF expression was assessed by ELISA. RGCs were labeled by neurons and RGC markers TUJ1 and BRN3A for quantitative and morphological analysis. Apoptosis was detected using PI/Hoechst staining and TUNEL assay and quantified by ImageJ. RESULTS: Cell proliferation and migration in HRMECs were significantly higher in the 25 mM glucose-treated group (p < 0.001) but lower in the 50 mM and 100 mM groups (p < 0.001). The permeability and the apoptotic index in HRMECs were statistically higher in the 25 mM, 50 mM, and 100 mM groups (p < 0.05). The tube formation assay found that all the parameters were significantly higher in the 25 mM and 50 mM groups (p < 0.001) concomitant with the elevated VEGFA expression in HRMECs (p = 0.016). Cell viability was significantly lower in the 50 mM, 100 mM, and 150 mM groups in RGCs (p 50mM = 0.013, p 100mM = 0.019, and p 150mM = 0.002). Apoptosis was significantly elevated, but the proportion of RGCs with neurite extension was significantly lower in the 50 mM, 100 mM, and 150 mM groups (p 50mM < 0.001, p 100mM < 0.001, and p 150mM < 0.001). CONCLUSIONS: We have optimized glucose concentrations to model diabetic retinal endothelial (25-50 mM) or neuronal (50-100 mM) dysfunction in vitro, which have a wide range of downstream applications.

Plant Chemistry Determines Host Preference and Performance of an Invasive Insect.

Understanding how host plant chemistry affects invasive insects is crucial for determining the physiological mechanism of host use and predicting invasive insect outbreak and damage on hosts. Here, we examined the effects of plant nutrition and defensive chemicals on host preference and performance of adults and larvae of the invasive potato tuberworm, Phthorimaea operculella (Zeller; Lepidoptera: Gelechiidae), on four native (Solanum tuberosum, Nicotiana tabacum, Datura stramonium, and Solanum lycopersicum) and three new (Solanum melongena, Physalis alkekengi, and Lycium barbarum) host plants. We found that adults preferred to oviposit on S. tuberosum and N. tabacum leaves and the soil around these native host plants over other hosts. Larvae performed well on S. tuberosum and N. tabacum, reaching higher pupa weight and having better survival. Larvae performed poorly on S. melongena, S. lycopersicum, P. alkekengi, D. stramonium, and L. barbarum, with lower pupa weight and lower survival. Solanum tuberosum and N. tabacum had higher leaf soluble proteins than other plants and lower leaf total phenolics than S. lycopersicum, D. stramonium, and L. barbarum. Moreover, carbon content and soluble protein were positively associated with larval survival, while defensive traits (lignin and total phenolics) negatively affected larval survival. These findings provide insights into understanding of biochemical mechanisms of interactions between invasive insects and host plants, indicating the importance of considering plant chemistry when assessing invasive insect host use and damage.

Histone deacetylase inhibitors differentially regulate c-Myc expression in retinoblastoma cells.

Retinoblastoma (RB) is the most prevalent childhood intraocular cancer type. Previous studies have demonstrated that c-myc (a proto-oncogene) is associated with tumorigenesis. However, at present, the influence of the expression profile and bioactivity of c-Myc on RB occurrence and progression is yet to be characterised. Notably, the present study demonstrated that c-myc is downregulated in the RB cell line WERI-Rb1. However, treatment with the histone deacetylase (HDAC) inhibitor trichostatin A (TSA) was revealed to significantly upregulate the expression of c-Myc mRNA and protein in WERI-Rb1 cells. Moreover, TSA increased the activity of the c-myc promoter in WERI-Rb1 cells, and the expression of c-Myc was also regulated by other HDAC inhibitors, including vorinostat (SAHA), valproic acid sodium salt (VPA) and entinostat. Notably, although c-myc was silenced in the Y79 cell line, the HDAC inhibitor TSA did not induce upregulation of mRNA and protein in Y79 cells. By contrast, certain HDAC inhibitors (TSA, VPA and SAHA) were discovered to significantly decrease the activity of the c-myc promoter in Y79 cells. Furthermore, the current data indicated that exogenous c-myc expression has a mild inhibitory effect on WERI-Rb1 and Y79 cell viability. Therefore, the present study revealed novel insights into the expression mechanism and bioactivity of c-Myc in RB cells.

Tetramethylpyrazine in a Murine Alkali-Burn Model Blocks NFκB/NRF-1/CXCR4-Signaling-Induced Corneal Neovascularization.

Purpose: Tetramethylpyrazine (TMP) is the active ingredient extracted from the Chinese herb Chuanxiong. The purpose of our study was to identify the mechanism of therapeutic TMP suppression of pathologic chemokine receptor 4 (CXCR4) transcription. Methods: C57BL/6J mice with alkali-burned corneas were treated with either TMP eye drops (1.5 mg/mL) or PBS. Corneal neovascularization (CNV) was measured and a clinical assessment was made by slit lamp microscopy. Expression of CXCR4 and the transcription factors nuclear respiratory factor-1 (NRF-1), nuclear factor kappa B (NFκB), forkhead box C1, and yin yang 1 were tracked by real-time RT-PCR and immunofluorescence staining of murine corneas. Western blot, real-time PCR, and immunofluorescence evaluated expression of related genes in human umbilical vein endothelial cells (HUVECs) after 200-µmol/L TMP treatment. In addition, plasmid transfection and chromatin immunoprecipitation assays elucidated the relationship among NRF-1, NFκB, and CXCR4. Results: Corneas treated with TMP had smaller areas of neovascularization and scored better in clinical assessments. Injured corneas showed significantly elevated expressions of NRF-1, NFκB, and CXCR4 that were normalized in vivo by TMP treatment. Similarly, in HUVECs in vitro, TMP decreased expression of NRF-1, NFκB, and CXCR4. Overexpression of NFκB or NRF-1 raised the expression of CXCR4 in HUVECs, but not synergistically. Chromatin immunoprecipitation assays detected only NRF-1 bound to the CXCR4 promoter region, suggesting NFκB controls CXCR4 expression by upregulating NRF-1. Together, our data suggest TMP downregulates CXCR4 by repressing NRF-1 expression in CNV, likely indirectly by downregulating NFκB. Conclusions: Our results implicate a novel mechanism wherein TMP inhibits neovascularization via an NFκB/NRF-1/CXCR4 circuit.

Brca1 Is Upregulated by 5-Aza-CdR and Promotes DNA Repair and Cell Survival, and Inhibits Neurite Outgrowth in Rat Retinal Neurons.

Previous studies have reported that Brca1 acts as a “hinge” in the development of the central nervous system (CNS). However, the precise role of Brca1 in rat retinal neurons remains unclear. Here, we found that Brca1 is developmentally downregulated and silenced in adult retina. Brca1 was upregulated in rat primary retinal neurons by 5-Aza-2′-deoxycytidine (5-Aza-CdR) treatment. Moreover, the upregulation of Brca1 by both 5-Aza-CdR and transgenic Brca1 promoted genomic stability and improved cell viability following exposure to ionizing radiation (IR). Furthermore, transgenic Brca1 significantly inhibited neurite outgrowth of retinal neurons, which implicates that Brca1 silencing promotes cell differentiation and determines neuronal morphology. Taken together, our results reveal a biological function of Brca1 in retinal development.

Discordant mRNA and protein expression of CXCR4 under in vitro CoCl2-induced hypoxic conditions.

Cobalt chloride (CoCl2) has long been accepted as a suitable in vitro hypoxia-mimetic agent. The gene CXCR4, which encodes a chemokine receptor, plays a key role in hypoxic retinal disease. Here, we investigated the mRNA and protein expression of CXCR4 in WERI-Rb1 retinoblastoma cells and human umbilical vein endothelial cells (HUVECs) under CoCl2-induced hypoxic conditions, by means of real-time PCR and western blot. We found that CoCl2-induced hypoxia profoundly increased CXCR4 expression at the mRNA level, but not at the protein level, at 12, 24, 48 and 72 h in these cells. Interestingly, this result differed from observations of 1% O2 hypoxic conditions. Additionally, luciferase assays demonstrated that CoCl2-induced hypoxia significantly increased transcription at the CXCR4 promoter. In order to compare our in vitro findings with the effects of hypoxia in vivo, an OIR (Oxygen-induced retinopathy) rat model was constructed. However, both CXCR4 mRNA and protein levels in OIR rats were significantly increased compared to controls. Thus taken together, our findings suggest that the relationship between CXCR4 mRNA and protein expression is not strictly linear under in vitro CoCl2-induced hypoxic conditions. through comparative in vitro and in vivo experiments, this study implies that CoCl2 is an imperfect simulation of hypoxia in retinal disease.