SUMMARY Visualizing dynamics of kinase activity in living animals is essential for mechanistic understanding of cell and developmental biology. We describe GFP-based kinase reporters that phase-separate upon kinase activation via multivalent protein-protein interactions, forming intensively fluorescent droplets. Called SPARK (separation of phases-based activity reporter of kinase), these reporters have large dynamic range (fluorescence change), high brightness, fast kinetics, and are reversible. The SPARK-based protein kinase A (PKA) reporter reveals oscillatory dynamics of PKA activities upon G protein-coupled receptor activation. The SPARK-based extracellular signal-regulated kinase (ERK) reporter unveils transient dynamics of ERK activity during tracheal metamorphosis in live Drosophila. Because of intensive brightness and simple signal pattern, SPARKs allow easy examination of kinase signaling in living animals in a qualitative way. The modular design of SPARK will facilitate development of reporters of other kinases.

The use of angiotensin receptor blockers (ARBs) correlates with reduced onset and progression of Alzheimer's disease (AD). The mechanism depicting how ARBs such as losartan restore cerebrovascular and cognitive deficits in AD is unknown. Here, we propose a mechanism underlying losartan's benefits by selectively blocking the effects of angiotensin IV (AngIV) at its receptor (AT4R) with divalinal in mice overexpressing the AD-related Swedish and Indiana mutations of the human amyloid precursor protein (APP mice) and WT mice. Young (3-month-old) mice were treated with losartan (ϳ10 mg/kg/d, 4 months), followed by intracerebroventricular administration of vehicle or divalinal in the final month of treatment. Spatial learning and memory were assessed using Morris water mazes at 3 and 4 months of losartan treatment. Cerebrovascular reactivity and whisker-evoked neurovascular coupling responses were measured at end point (ϳ7 months of age), together with biomarkers related to neuronal and vascular oxidative stress (superoxide dismutase-2), neuroinflammation (astroglial and microglial activation), neurogenesis (BrdU-labeled newborn cells), and amyloidosis [soluble amyloid-␤ (A␤) species and A␤ plaque load]. Divalinal countered losartan's capacity to rescue spatial learning and memory and blocked losartan's benefits on dilatory function and baseline nitric oxide bioavailability. Divalinal reverted losartan's anti-inflammatory effects, but failed to modify losartan-mediated reductions in oxidative stress. Neither losartan nor divalinal affected arterial blood pressure or significantly altered the amyloid pathology in APP mice. Our findings identify activation of the AngIV/AT4R cascade as the underlying mechanism in losartan's benefits and a target that could restore A␤-related cognitive and cerebrovascular deficits in AD.

Akt-mediated signaling plays an important role in blood vascular development. In this study , we investigated the role of Akt in lymphatic growth using Akt-deficient mice. First , we found that lymphangiogenesis occurred in Akt1 ؊/؊ , Akt2 ؊/؊ , and Akt3 ؊/؊ mice. However , both the diameter and endothelial cell number of lymphatic capillaries were significantly less in Akt1 ؊/؊ mice than in wild-type control mice , whereas there was only a slight change in Akt2 ؊/؊ and Akt3 ؊/؊ mice. Second , valves present in the small collecting lymphatics in the superficial dermal layer of the ear skin were rarely observed in Akt1 ؊/؊ mice , although these valves could be detected in the large collecting lymphatics in the deep layer of the skin tissues. A fluorescence microlymphangiography assay showed that the skin lymphatic network in Akt1 ؊/؊ mice was functional but abnormal as shown by fluorescein isothiocyanate-dextran draining. There was an uncharacteristic enlargement of collecting lymphatic vessels , and further analysis showed that smooth muscle cell coverage of collecting lymphatic vessels became much more sparse in Akt1-deficient mice than in wild-type control animals. Finally , we showed that lymphatic vessels were detected in compound Akt-null mice and that lymphangiogenesis could be induced by vascular endothelial growth factor-C delivered via adenoviral vectors in adult mice lacking Akt1. These results indicate that despite the compensatory roles of other Akt isoforms, Akt1 is more critically required during lymphatic development.

Universities and medical schools in China are faced with an ongoing shortage of cadavers for education and research because of insufficient numbers of cadaver donations. This article will examine the main obstacles to cadaver donation in the Chinese culture. These include superstitious traditional views about the body, a lack of legislation regulating donations, and a deficiency of effective channels for cadaver donations. Cadaver dissection has always been the most important method of teaching anatomy to medical students. Today, ethics courses have also become essential to a complete medical education. Contemporary physicians need to be equipped to navigate the myriad of moral and ethical issues inherent to modern medicine. In China, cadaver donations lag behind those in other countries, threatening to create valid disadvantages in medical education. New legislation and public education are necessary to remove cultural barriers and change Chinese views on cadaver donation. For this reason, the Department of Human Anatomy at Nanjing Medical University has established the "Educational Center for Medical Ethics." The goal of the Center is to promote proper respect for cadavers used for medical research and education, cherish the human lives the cadavers represent, and gain the trust of potential donors.

Adipose tissues are classified into white adipose tissue (WAT) and brown adipose tissue (BAT). WAT is responsible for energy storage, and malfunction is associated with obesity. BAT, on the contrary, consumes fat to generate heat through uncoupling mitochondrial respiration and is important in body weight control. Vascular endothelial growth factor (VEGF)-A is the founding member of the VEGF family and has been found highly expressed in adipose tissue. A genetic mouse model of an inducible VEGF (VEGF-A) repression system was used to study VEGF-regulated energy metabolism in WAT. VEGF-repressed mice demonstrated lower food efficiency, lower body weight, and resistance to high-fat diet-induced obesity. Repression of VEGF expression caused morphological and molecular changes in adipose tissues. VEGF repression induced brown-like adipocyte development in WAT, up-regulation of BAT-specific genes including PRDM16, GATA-1, BMP-7, CIDEA, and UCP-1 and down-regulation of leptin, a WAT-specific gene. VEGF repression up-regulated expression of VEGF-B and its downstream fatty acid transport proteins. Relative levels of VEGF/VEGF-B may be important switches in energy metabolism and of pharmaceutical significances.

The use of human tissue is critical for gross anatomy education in the health professions. Chinese medical colleges have faced a shortage of anatomical specimens over the past decade. While body donation plays an important role in overcoming this gap, this practice has only recently been introduced in China, and the donation rate is relatively low and fraught with a number of difficulties. In the past, traditional Chinese culture focused on preserving the human body intact, which often limited body donation. In recent years, the public has become more open toward body donation. At Nanjing Medical University, only 20 bodies were donated in 2001. After the university became involved in an organized body donation program, this number increased to 70 donated bodies per year (2007 to 2012). This article describes and reviews Chinese medical colleges as a special case study among body donation programs, particularly in terms of the multiple responsibilities and roles that such institutions must assume in the course of adopting these programs. Medical colleges in China must serve as advocates, coordinators, builders, managers, educators, and beneficiaries in undertaking body donation programs. It is important for medical colleges to recognize these pluripotent roles and educate the public in order to promote body donation programs. This case study may also effectively guide and encourage Chinese medical colleges in refining their own body donation programs in the future.

Objective-The genetic program underlying lymphatic development is still incompletely understood. This study aims to dissect the role of receptor tyrosine kinase with immunoglobulin-like and EGF (epidermal growth factor)-like domains 1 (Tie1) and Tie2 in lymphatic formation using genetically modified mouse models. Approach and Results-We generated conditional knockout mouse models targeting Tie1, Tie2, and angiopoietin-2 in this study. Tie1 ΔICD/ΔICD mice, with its intracellular domain targeted, appeared normal at E10.5 but displayed subcutaneous edema by E13.5. Lymph sac formation occurred in Tie1 ΔICD/ΔICD mice, but they had defects with the remodeling of primary lymphatic network to form collecting vessels and valvulogenesis. Consistently, induced deletion of Tie1-ICD postnatally using a ubiquitous Cre deleter led to abnormal lymphangiogenesis and valve formation in Tie1-ICD iUCKO/− mice. In comparison with the lymphatic phenotype of Tie1 mutants, we found that the diameter of lymphatic capillaries was significantly less in mice deficient of angiopoietin-2, besides the disruption of collecting lymphatic vessel formation as previously reported. There was also no lymphedema observed in Ang2 −/− mice during embryonic development, which differs from that of Tie1 ΔICD/ΔICD mice. We further investigated whether Tie1 exerted its function via Tie2 during lymphatic development. To our surprise, genetic deletion of Tie2 (Tie2 Key Words: knockout mice ◼ lymphatic abnormality ◼ lymphatic vessel ◼ Tie-1 receptor tyrosine kinase ◼ Tie-2 receptor tyrosine kinase ◼ valve

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