Calpain inhibitors reduce retinal hypoxia in ischemic retinopathy by improving neovascular architecture and functional perfusion

AbstractIn ischemic retinopathies, underlying hypoxia drives abnormal neovascularization that damages retina and causes blindness. The abnormal neovasculature is tortuous and leaky and fails to alleviate hypoxia, resulting in more pathological neovascularization and retinal damage. With an established model of ischemic retinopathy we found that calpain inhibitors, when administered in moderation, reduced architectural abnormalities, reduced vascular leakage, and most importantly reduced retinal hypoxia. Mechanistically, these calpain inhibitors improved stability and organization of the actin cytoskeleton in retinal endothelial cells undergoing capillary morphogenesis in vitro, and they similarly improved organization of actin cables within new blood vessels in vivo. Hypoxia induced calpain activity in retinal endothelial cells and severely disrupted the actin cytoskeleton, whereas calpain inhibitors preserved actin cables under hypoxic conditions. Collectively, these findings support the hypothesis that hyper-activation of calpains by hypoxia contributes to disruption of the retinal endothelial cell cytoskeleton, resulting in formation of neovessels that are defective both architecturally and functionally. Modest suppression of calpain activity with calpain inhibitors restores cytoskeletal architecture and promotes formation of a functional neovasculature, thereby reducing underlying hypoxia. In sharp contrast to “anti-angiogenesis” strategies that cannot restore normoxia and may aggravate hypoxia, the therapeutic strategy described here does not inhibit neovascularization. Instead, by improving the function of neovascularization to reduce underlying hypoxia, moderate calpain inhibition offers a method for alleviating retinal ischemia, thereby suggesting a new treatment paradigm based on improvement rather than inhibition of new blood vessel growth

Moderation of Calpain Activity Promotes Neovascular Integration and Lumen Formation during VEGF-Induced Pathological Angiogenesis

Successful neovascularization requires that sprouting endothelial cells (ECs) integrate to form new vascular networks. However, architecturally defective, poorly integrated vessels with blind ends are typical of pathological angiogenesis induced by vascular endothelial growth factor-A (VEGF), thereby limiting the utility of VEGF for therapeutic angiogenesis and aggravating ischemia-related pathologies. Here we investigated the possibility that over-exuberant calpain activity is responsible for aberrant VEGF neovessel architecture and integration. Calpains are a family of intracellular calcium-dependent, non-lysosomal cysteine proteases that regulate cellular functions through proteolysis of numerous substrates.In a mouse skin model of VEGF-driven angiogenesis, retroviral transduction with dominant-negative (DN) calpain-I promoted neovessel integration and lumen formation, reduced blind ends, and improved vascular perfusion. Moderate doses of calpain inhibitor-I improved VEGF-driven angiogenesis similarly to DN calpain-I. Conversely, retroviral transduction with wild-type (WT) calpain-I abolished neovessel integration and lumen formation. In vitro, moderate suppression of calpain activity with DN calpain-I or calpain inhibitor-I increased the microtubule-stabilizing protein tau in endothelial cells (ECs), increased the average length of microtubules, increased actin cable length, and increased the interconnectivity of vascular cords. Conversely, WT calpain-I diminished tau, collapsed microtubules, disrupted actin cables, and inhibited integration of cord networks. Consistent with the critical importance of microtubules for vascular network integration, the microtubule-stabilizing agent taxol supported vascular cord integration whereas microtubule dissolution with nocodazole collapsed cord networks.These findings implicate VEGF-induction of calpain activity and impairment of cytoskeletal dynamics in the failure of VEGF-induced neovessels to form and integrate properly. Accordingly, calpain represents an important target for rectifying key vascular defects associated with pathological angiogenesis and for improving therapeutic angiogenesis with VEGF

FIELD TEST ON CLEANING OF OIL POLLUTION ON NHATRANG BEACH OF VIETNAM

Joint Research on Environmental Science and Technology for the Eart

Information searching behaviors among Vietnamese students during first wave of the COVID-19 pandemic

This study aims to describe the COVID-19 related information searching behaviors and the relationship between those behaviors and the satisfaction with the COVID-19 related information searched on the Internet among university students during first wave of the COVID-19 pandemic in Vietnam

Effects of metformin on retinoblastoma growth in vitro and in vivo

Recent studies suggest that the anti-diabetic drug metformin may reduce the risk of cancer and have anti-proliferative effects for some but not all cancers. In this study, we examined the effects of metformin on human retinoblastoma cell proliferation in vitro and in vivo. Two different human retinoblastoma cell lines (Y79, WERI) were treated with metformin in vitro and xenografts of Y79 cells were established in nu/nu immune-deficient mice and used to assess the effects of pharmacological levels of metformin in vivo. Metformin inhibited proliferation of the retinoblastoma cells in vitro. Similar to other studies, high concentrations of metformin (mM) blocked the cell cycle in G0–G1, indicated by a strong decrease of G1 cyclins, especially cyclin D, cyclin-dependent kinases (4 and 6), and flow cytometry assessment of the cell cycle. This was associated with activation of AMPK, inhibition of the mTOR pathways and autophagy marker LC3B. However, metformin failed to suppress growth of xenografted tumors of Y79 human retinoblastoma cells in nu/nu mice, even when treated with a maximally tolerated dose level achieved in human patients. In conclusion, suprapharmacological levels (mM) of metformin, well above those tolerated in vivo, inhibited the proliferation of retinoblastoma cells in vitro. However, physiological levels of metformin, such as seen in the clinical setting, did not affect the growth of retinoblastoma cells in vitro or in vivo. This suggests that the potential beneficial effects of metformin seen in epidemiological studies may be limited to specific tumor types or be related to indirect effects/mechanisms not observed under acute laboratory conditions

NGHIÊN CỨU VI SINH VẬT SỐNG CÙNG MỘT SỐ LOÀI SAN HÔ CỨNG TẠI HANG RÁI NINH THUẬN BẰNG PHƯƠNG PHÁP NHUỘM HUỲNH QUANG KẾT HỢP NUÔI CẤY TỚI HẠN

Coral associated bacteria and their host are currently one of the interested issues for research and scientists worldwide. The densities of zooxanthellae and bacteria associated with three most prevalent species Acropora hyacinthus, Acropora muricata and Acropora robusta in Hang Rai, Ninh Thuan was evaluated over time by staining with SYBR Gold and direct counting with epifluorescence method. The most dominant bacteria were isolated by culture dependent method. The densities of zooxanthellae and bacteria ranged from 0.39–1.83×107 cell/g, and 0.83–2.52×108 cell/g, respectively. Bacterial density in the 3 months was significantly different compared to the density of the bacteria in ambient water. Total heterotrophic bacteria, comma shaped bacteria and bacillus form showed negatively correlated with pH, PO4, while zooxanthellae showed no correlation with all factors.Rạn san hô trên toàn thế đang đối mặt với sự huỷ diệt nghiêm trọng, một trong những nguyên nhân chính là do vi khuẩn gây bệnh và những tác động của môi trường. Chính vì vậy, nghiên cứu về hệ vi khuẩn sống cùng san hô và mối tương quan giừa vi khuẩn, san hô và các yếu tố môi trường là quan trọng và cấp thiết. Trong nghiên cứu này, mật độ vi tảo Symbiodinium sp., vi khuẩn sống cùng 3 loài san hô cứng Acropora hyacinthus, Acropora muricata và Acropora robusta phổ biến tại Ninh Thuận được đánh giá vào các thời điểm trước, trong, và sau khi san hô bị tẩy trắng bằng phương pháp đếm huỳnh quang và pha loãng tới hạn. Kết quả cho thấy mật độ tảo Symbiodinium khác nhau có ý nghĩa thống kê (dao động 0,39-1,83x107tb/g) ở các loài san hô khác nhau. Tuy nhiên, mật độ tảo cộng sinh không có khác biệt lớn giữa các tháng nghiên cứu. Mật độ vi khuẩn dao động từ 0,83-2,52x108tb/g và có sự sai khác có ý nghĩa thống kê không chỉ giữa các loài san hô mà còn ở các thời điểm trước trong và sau tẩy trắng. Tổng vi khuẩn, phẩy khuẩn và trực khuẩn có tương quan nghịch và có ý nghĩa về mặt thống kê với chỉ số pH và hàm lượng PO4. Ngược lại, mật độ tảo hoàn toàn không tương quan với các yếu tố môi trường

Issues with the Specificity of Immunological Reagents for NLRP3: Implications for Age-related Macular Degeneration

Contradictory data have been presented regarding the implication of the NACHT, LRR and PYD domains-containing protein 3 (NLRP3) inflammasome in age-related macular degeneration (AMD), the leading cause of vision loss in the Western world. Recognizing that antibody specificity may explain this discrepancy and in line with recent National Institutes of Health (NIH) guidelines requiring authentication of key biological resources, the specificity of anti-NLRP3 antibodies was assessed to elucidate whether non-immune RPE cells express NLRP3. Using validated resources, NLRP3 was not detected in human primary or human established RPE cell lines under multiple inflammasome-priming conditions, including purported NLRP3 stimuli in RPE such as DICER1 deletion and Alu RNA transfection. Furthermore, NLRP3 was below detection limits in ex vivo macular RPE from AMD patients, as well as in human induced pluripotent stem cell (hiPSC)-derived RPE from patients with overactive NLRP3 syndrome (Chronic infantile neurologic cutaneous and articulate, CINCA syndrome). Evidence presented in this study provides new data regarding the interpretation of published results reporting NLRP3 expression and upregulation in RPE and addresses the role that this inflammasome plays in AMD pathogenesis