Tankyrase inhibition interferes with junction remodeling, induces leakiness, and disturbs YAP1/TAZ signaling in the endothelium.

Tankyrase inhibitors are increasingly considered for therapeutic use in malignancies that are characterized by high intrinsic ß-catenin activity. However, how tankyrase inhibition affects the endothelium after systemic application remains poorly understood. In this study, we aimed to investigate how the tankyrase inhibitor XAV939 affects endothelial cell function and the underlying mechanism involved. Endothelial cell function was analyzed using sprouting angiogenesis, endothelial cell migration, junctional dynamics, and permeability using human umbilical vein endothelial cells (HUVEC) and explanted mouse retina. Underlying signaling was studied using western blot, immunofluorescence, and qPCR in HUVEC in addition to luciferase reporter gene assays in human embryonic kidney cells. XAV939 treatment leads to altered junctional dynamics and permeability as well as impaired endothelial migration. Mechanistically, XAV939 increased stability of the angiomotin-like proteins 1 and 2, which impedes the nuclear translocation of YAP1/TAZ and consequently suppresses TEAD-mediated transcription. Intriguingly, XAV939 disrupts adherens junctions by inducing RhoA-Rho dependent kinase (ROCK)-mediated F-actin bundling, whereas disruption of F-actin bundling through the ROCK inhibitor H1152 restores endothelial cell function. Unexpectedly, this was accompanied by an increase in nuclear TAZ and TEAD-mediated transcription, suggesting differential regulation of YAP1 and TAZ by the actin cytoskeleton in endothelial cells. In conclusion, our findings elucidate the complex relationship between the actin cytoskeleton, YAP1/TAZ signaling, and endothelial cell function and how tankyrase inhibition disturbs this well-balanced signaling.

Sufficient Cav-1 levels in the endothelium are critical for the maintenance of the neurovascular unit in the retina.

BACKGROUND: Caveolin-1 (Cav-1) is a pivotal protein in the plasma membrane. Studies on homozygous Cav-1 deficient mice revealed that Cav-1 is essential for endothelial function and angiogenesis in the retina. However, whether a reduction in Cav-1 content hampers the neurovascular unit (NVU) in the retina is unclear. Thus, this study examines the NVU in the retinas of heterozygous Cav-1 deficient (Cav-1+/-) mice and analyzes possible underlying mechanisms. METHODS: The vascular, glial and neuronal components in the retina were evaluated using retinal morphometry, whole mount retinal immunofluorescence staining, histological analysis and optical coherence tomography. In addition, immunoblotting and immunofluorescence staining, subcellular fractionation, biotin labeling of cell surface proteins, and proximity ligation assay were employed to detect expression and localization of proteins in the retina or endothelial cells (ECs) upon knockdown of Cav-1 with Cav-1 siRNA. RESULTS: Cav-1+/- retinas showed a significant reduction in pericyte coverage along with an increase in acellular capillaries compared to controls at 8 months of age, but not at 1 month. A significant loss and obvious morphological abnormalities of smooth muscle cells were observed in 8-month-old Cav-1+/- retinal arterioles. Macroglial and microglial cells were activated in the Cav-1+/- retinas. A transient significant delay in retinal angiogenesis was detected in Cav-1+/- retinas at p5, which was however no longer detectable at p10. The Cav-1+/- retinas displayed increased vascular permeability and a notable reduction in VEGFR2 content at 8 months. In vitro, siRNA-mediated knockdown experiments in ECs revealed that the loss of Cav-1 in ECs resulted in decreased levels of VEGFR2, VE-Cadherin and their interaction at the plasma membrane as well. CONCLUSION: Our results indicate that a sufficient Cav-1 level over 50% of its normal abundance is vital for the proper localization of VEGFR2 and VE-cadherin, likely in a complex, at the plasma membrane, which is essential for the maintenance of normal NVU in the retina.

Acute Effects of Breakfast Fruits Meal Sequence and Postprandial Exercise on the Blood Glucose Level and DPP4 Activity among Type 2 Diabetes Mellitus Patients: A Pilot Study.

Objectives: Type 2 diabetes mellitus (T2DM) is a major global public health issue. Diet and physical exercise are modifiable factors that influence the glycaemic status of patients with T2DM. We aimed to investigate the acute effects of breakfast fruits meal sequence and postprandial exercise on the blood glucose level and dipeptidyl peptidase 4 (DPP4) activity among type 2 diabetes mellitus patients. Methods: A randomized pilot study recruited patients with T2DM who attended two primary health care centres in Tasikmadu District, Karanganyar Regency, and Kartasura District, Sukoharjo Regency, Central Java, Indonesia, from July to October 2016. Eligible patients (4 men and 32 women) were randomly divided into four treatment groups. Venous blood samples were analyzed for fasting and one-hour postprandial blood glucose (FBG and 1 h PPG) levels and DPP4 activity. Blood glucose levels were measured using a routine hexokinase method, and serum DPP4 activity was determined spectrophotometrically after incubation with the Gly-Pro-p-nitroanilide substrate. Results: Fruits last meal decreased FBG level whilst fruits first meal did not significantly decrease 1 h PPG level. Both treatments had no acute effects on DPP4 activity but the addition of postprandial exercise helped lower DPP4 activity. Fruit last and first meals showed significant opposite effects on mean changes of FBG level (p < 0.05). Conclusions: This preliminary report of fruits meal sequence is potentially involved in acute regulation of blood glucose levels and that it might be independent of DPP4 activity in Indonesian patients with T2DM. Moreover, postprandial exercise may be an important intervention for T2DM through the mediation of DPP4 but has no acute effects on the regulation of blood glucose levels. Further studies are required to investigate whether or not different types of fruits and longer treatment intervals can affect blood glucose levels and DPP4 activity differently. This study also gives an insight into the feasibility of conducting food order modification with or without the combination of postprandial exercise in a primary health setting for our next studies.

Effects of Vegetables Consumption Before Carbohydrates on Blood Glucose and GLP-1 Levels Among Diabetic Patients in Indonesia.

Background: Type 2 Diabetes Mellitus (T2DM) is the prominent public health issue. Pharmacotherapy and diet modification should be integrated into T2DM management. Aims: To investigate the effects of vegetables consumption before carbohydrates on blood glucose and GLP-1 levels in T2DM patients. Methods: A non-randomized quasi experimental study was conducted to recruit T2DM patients who attended at the Gatot Soebroto Central Army Hospital, Jakarta, Indonesia from April to May 2016. The Lemeshow's formula was used to determine sample size. A total of 12 non-diabetic and 24 diabetic patients were participated in our study. Glucose levels were measured using a routine hexokinase method while serum GLP-1 levels were determined using the ELISA. The student t-test was used to compare two groups with parametric data. The significant difference was at P < 0.05. Results: Our data showed that T2DM patients who consumed vegetables before carbohydrates, had relatively stable glucose levels at 0, 60 and 120 mins (164.25 ± 86.89 vs 183.5 ± 55.96 vs 167.83 ± 65.53, P = 0.163) and stay lowered within the normal range compared to T2DM patients who consumed vegetables after carbohydrates (165.08 ± 67.89 vs 241.92 ± 68.03 vs 204.92 ± 81.76, P = 0.022). Additionally, GLP-1 levels remained stable after 60 and 120 min at day 1 (P = 0.816) and day 3 (P = 0.955). Conclusions: Vegetables consumption before carbohydrate is a promising and simple method of diabetes diet for maintaining blood glucose and GLP-1 levels and preventing from vascular complication.