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The purpose of crowd counting is to estimate the number of pedestrians in crowd images. Crowd counting or density estimation is an extremely challenging task in computer vision, due to large scale variations and dense scene. Current methods solve these issues by compounding multi-scale Convolutional Neural Network with different receptive fields. In this paper, a novel end-to-end architecture based on Multi-Scale Adversarial Convolutional Neural Network (MSA-CNN) is proposed to generate crowd density and estimate the amount of crowd. Firstly, a multi-scale network is used to extract the globally relevant features in the crowd image, and then fractionally-strided convolutional layers are designed for up-sampling the output to recover the loss of crucial details caused by the earlier max pooling layers. An adversarial loss is directly employed to shrink the estimated value into the realistic subspace to reduce the blurring effect of density estimation. Joint training is performed in an end-to-end fashion using a combination of Adversarial loss and Euclidean loss. The two losses are integrated via a joint training scheme to improve density estimation performance.We conduct some extensive experiments on available datasets to show the significant improvements and supremacy of the proposed approach over the available state-of-the-art approaches.

Diabetes increases oxidant stress and doubles the risk of dying after myocardial infarction, but the mechanisms underlying increased mortality are unknown. Mice with streptozotocin-induced diabetes developed profound heart rate slowing [...]

The E3 ubiquitin ligase NEDD4-2 (encoded by the Nedd4L gene) regulates the amiloride-sensitive epithelial [Na.sup.+] channel (ENaC/SCNN1) to mediate [Na.sup.+] homeostasis. Mutations in the human β/γENaC subunits that block NEDD4-2 binding or constitutive ablation of exons 6-8 of Nedd4L in mice both result in salt-sensitive hyper-tension and elevated ENaC activity (Liddle syndrome). To determine the role of renal tubular NEDD4-2 in adult mice, we generated tetracycline-inducible, nephron-specific Nedd4L KO mice. Under standard and high-[Na.sup.+] diets, conditional KO mice displayed decreased plasma aldosterone but normal [Na.sup.+]/[K.sup.+] balance. Under a high-[Na.sup.+] diet, KO mice exhibited hypercalciuria and increased blood pressure, which were reversed by thiazide treatment. Protein expression of βENaC, γENaC, the renal outer medullary [K.sup.+] channel (ROMK), and total and phosphorylated thiazide-sensitive [Na.sup.+][Cl.sup.-] cotransporter (NCC) levels were increased in KO kidneys. Unexpectedly, Scnn1a mRNA, which encodes the αENaC subunit, was reduced and proteolytic cleavage of αENaC decreased. Taken together, these results demonstrate that loss of NEDD4-2 in adult renal tubules causes a new form of mild, salt-sensitive hypertension without hyperkalemia that is characterized by upregulation of NCC, elevation of β/γENaC, but not αENaC, and a normal [Na.sup.+]/[K.sup.+] balance maintained by downregulation of ENaC activity and upregulation of ROMK., Introduction Hypertension is a major risk factor for stroke, myocardial infarction, and heart and kidney failure. The postmacula densa segments of the nephron, namely the distal convoluted tubule (DCT), the [...]

We report a mouse model of multiple osteochondromas (MO), an autosomal dominant disease in humans, also known as multiple hereditary exostoses (MHE or HME) and characterized by the formation of cartilage-capped osseous growths projecting from the metaphyses of endochondral bones. The pathogenesis of these osteochondromas has remained unclear. Mice heterozygous for Ext1 or Ext2, modeling the human genotypes that cause MO, occasionally develop solitary osteochondroma-like structures on ribs [Lin et al. (2000) Dev Biol 224(2):299-311; Stickens et al. (2005) Development 132(22):5055-5068]. Rather than model the germ-line genotype, we modeled the chimeric tissue genotype of somatic loss of heterozygosity (LOH), by conditionally inactivating Ext1 via head-to-head loxP sites and temporally controlled Cre-recombinase in chondrocytes. These mice faithfully recapitulate the human phenotype of multiple metaphyseal osteochondromas. We also confirm homozygous disruption of Ext1 in osteochondroma chondrocytes and their origin in proliferating physeal chondrocytes. These results explain prior modeling failures with the necessity for somatic LOH in a developmentally regulated cell type. osteochondroma | exostosis | loss of heterozygosity | growth plate | bone doi/10.1073/pnas.0910875107

The regulation of metal ion transport within neurons is critical for normal brain function. Of particular importance is the regulation of redox metals such as iron (Fe), where excess levels can contribute to oxidative stress and protein aggregation, leading to neuronal death. The divalent metal transporter 1 (DMT1) plays a central role in the regulation of Fe as well as other metals; hence, failure of DMT1 regulation is linked to human brain pathology. However, it remains unclear how DMT1 is regulated in the brain. Here, we show that DMT1 is regulated by Ndfip1 (Nedd4 family-interacting protein 1), an adaptor protein that recruits E3 ligases to ubiquitinate target proteins. Using human neurons we show the Ndfip1 is upregulated and binds to DMT1 in response to Fe and cobalt (Co) exposure. This interaction results in the ubiquitination and degradation of DMT1, resulting in reduced metal entry. Induction of Ndfip1 expression protects neurons from metal toxicity, and removal of Ndfip1 by shRNAi results in hypersensitivity to metals. We identify Nedd4-2 as an E3 ligase recruited by Ndfip1 for the ubiquitination of DMT1 within human neurons. Comparison of brains from [Ndfip1.sup.-/-] with [Ndfip1.sup.+/+] mice exposed to Fe reveals that [Ndfip1.sup.-/-] brains accumulate Fe within neurons. Together, this evidence suggests a critical role for Ndfip1 in regulating metal transport in human neurons. cobalt | iron | Nedd4-2 | ubiquitin

Nedd4-2 has been proposed to play a critical role in regulating epithelial [Na.sup.+] channel (ENaC) activity. Biochemical and overexpression experiments suggest that Nedd4-2 binds to the PY motifs of ENaC subunits via its WW domains, ubiquitinates them, and decreases their expression on the apical membrane. Phosphorylation of Nedd4-2 (for example by Sgkl) may regulate its binding to ENaC, and thus ENaC ubiquitination. These results suggest that the interaction between Nedd4-2 and ENaC may play a crucial role in [Na.sup.+] homeostasis and blood pressure (BP) regulation. To test these predictions in vivo, we generated Nedd4-2 null mice. The knockout mice had higher BP on a normal diet and a further increase in BP when on a high-salt diet. The hypertension was probably mediated by ENaC overactivity because 1) Nedd4-2 null mice had higher expression levels of all three ENaC subunits in kidney, but not of other [Na.sup.+] transporters; 2) the downregulation of ENaC function in colon was impaired; and 3) NaC1sensitive hypertension was substantially reduced in the presence of amiloride, a specific inhibitor of ENaC. Nedd4-2 null mice on a chronic high-salt diet showed cardiac hypertrophy and markedly depressed cardiac function. Overall, our results demonstrate that in vivo Nedd4-2 is a critical regulator of ENaC activity and BP. The absence of this gene is sufficient to produce salt-sensitive hypertension. This model provides an opportunity to further investigate mechanisms and consequences of this common disorder. ion channels; kidney; sodium channels

The intercalated disk protein Xin was originally discovered in chicken striated muscle and implicated in cardiac morphogenesis. In the mouse, there are two homologous genes, mXin[alpha] and toxin[beta]. The human homolog of mXin[alpha], Cmya1, maps to chromosomal region 3p21.2-21.3, near a dilated cardiomyopathy with conduction defect-2 locus. Here we report that mXin[alpha]-null mouse hearts are hypertrophied and exhibit fibrosis, indicative of cardiomyopathy. A significant upregulation of mXin[beta] likely provides partial compensation and accounts for the viability of the mXin[alpha]-null mice. Ultrastructural studies of mXin[alpha]-null mouse hearts reveal intercalated disk disruption and myofilament disarray. In mXin[alpha]-null mice, there is a significant decrease in the expression level of p120-catenin, [beta]-catenin, N-cadherin, and desmoplakin, which could compromise the integrity of the intercalated disks and functionally weaken adhesion, leading to cardiac defects. Additionally, altered localization and decreased expression of connexin 43 are observed in the mXin[alpha]-null mouse heart, which, together with previously observed abnormal electrophysiological properties of mXin[alpha]-deficient mouse ventricular myocytes, could potentially lead to conduction defects. Indeed, ECG recordings on isolated, perfused hearts (Langendorff preparations) show a significantly prolonged QT interval in mXin[alpha]-deficient hearts. Thus mXin[alpha] functions in regulating the hypertrophic response and maintaining the structural integrity of the intercalated disk in normal mice, likely through its association with adherens junctional components and actin cytoskeleton. The mXin[alpha]-knockout mouse line provides a novel model of cardiac hypertrophy and cardiomyopathy with conduction defects. Xin repeat proteins; N-cadherin; [beta]-catenin: p120-catenin; connexin 43

In mammals, the hormonal regulation of water homeostasis is mediated by the aquaporin-2 water channel (Aqp2) of the collecting duct (CD). Vasopressin induces redistribution of Aqp2 from intracellular vesicles to the apical membrane of CD principal cells, accompanied by increased water permeability. Mutations of AQP2 gene in humans cause both recessive and dominant nephrogenic diabetes insipidus (NDI), a disease in which the kidney is unable to concentrate urine in response to vasopressin. In this study, we generated a line of mice with the distal COOH-terminal tail of the Aqp2 deleted (Aqp[2.sup.[DELTA]230]), including the protein kinase A phosphorylation site (S256), but still retaining the putative apical localization signal (221-229) at the COOH-terminal. Mice heterozygous for the truncation appear normal. Homozygotes are viable to adulthood, with reduced urine concentrating capacity, increased urine output, decreased urine osmolality, and increased daily water consumption. Desmopressin increased urine osmolaiity in wild-type mice but had no effect on Aqp[2.sup.[DELTA]230/[DELTA]230] mice. Kidneys from affected mice showed CD and pelvis dilatation and papillary atrophy. By immunohistochemical and immunoblot analyses using antibody against the N[H.sub.2]-terminal region of the protein Aqp[2.sup.[DELTA]230/[DELTA]230] mice had a markedly reduced protein abundance. Expression of the truncated protein in MDCK cells was consistent with a small amount of functional expression but no stimulation. Thus we have generated a mouse model of NDI that may be useful in studying the physiology and potential therapy of this disease. mouse; aquaporin-2; diabetes insipidus; protein trafficking; protein phosphorylation doi:10.1152/ajprenal.00308.2006

The present studies were designed to determine whether mice heterozygous for deletion of [beta]-ENaC exhibited defects in [Na.sup.+]/[K.sup.+] transport and blood pressure regulation. In response to an acute KCI infusion, +/- mice developed higher serum [[K.sup.+]] and excreted only 40% of the [K.sup.+] excreted by +/+ mice. After 6 days on a low (0.01%)-[Na.sup.+] diet, the cumulative [Na.sup.+] excretion from days 3-6 was greater for +/mice. This low-[Na.sup.+] diet caused higher serum [[K.sup.+]] and lower [K.sup.+] excretion rates in +/- mice than in +/+ mice, but the rectal potential differences were not different. Analyses of mRNA from mice on this diet showed the expected ~50% reduction of [beta]-ENaC in kidney and colon of +/- mice. Unexpectedly, the level of [gamma]-ENaC mRNA was similarly reduced. NHE3 mRNA was ~30% higher in +/- mice whereas mRNA of the Na-K-2Cl cotransporter was not different. Also unexpectedly, the amount of [beta]-ENaC proteins was similar in both groups of mice but there was a reduction of one form of [gamma]-ENaC in +/- mice. These experiments demonstrate that mice heterozygous for [beta]-ENaC have a small but detectable defect in their ability to conserve [Na.sup.+] and a more readily apparent defect in the ability to secrete [K.sup.+]. sodium; homeostasis; heterozygote; low-[Na.sup.+] diet

Author(s): Margaret P. Price [1, 2]; Gary R. Lewin [1, 3]; Sabrina L. McIlwrath [3]; Chun Cheng [2]; Jinghui Xie [2]; Paul A. Heppenstall [3]; Cheryl L. Stucky [3]; Anne [...]

The epithelial [Na.sup.+] channel (ENaC) is composed of three homologous subunits: [Alpha], [Beta] and [Gamma]. We used gene targeting to disrupt the [Beta] subunit gene of ENaC in mice. The [Beta]ENaC-deficient mice showed normal prenatal development but died within 2 days after birth, most likely of hyperkalemia. In the -/- mice, we found an increased urine [Na.sup.+] concentration despite hyponatremia and a decreased urine [K.sup.+] concentration despite hyperkalemia. Moreover, serum aldosterone levels were increased. In contrast to [Alpha]ENaC-deficient mice, which die because of defective lung liquid clearance, neonatal [Beta]ENaC deficient mice did not die of respiratory failure and showed only a small increase in wet lung weight that had little, if any, adverse physiologic consequence. The results indicate that, in vivo, the [Beta] subunit is required for ENaC function in the renal collecting duct, but, in contrast to the [Alpha] subunit, the [Beta] subunit is not required for the transition from a liquid-filled to an air-filled lung. The phenotype of the [Beta]ENaC-deficient mice is similar to that of humans with pseudohypoaldosteronism type 1 and may provide a useful model to study the pathogenesis and treatment of this disorder.

We have used a 'plug and socket' targeting technique to generate a mouse model of [beta]0-thalassemia in which both the b] and b2 adult globin genes have been deleted. Mice homozygous for this deletion (Hbbl.sup.th-3/Hbb[sup.th-3]) die perinatally, similar to the most severe form of Cooley anemia in humans. Mice heterozygous for the deletion appear normal, but their hematologic indices show characteristics typical of severe thalassemia, including dramatically decreased hematocrit, hemoglobin, red blood cell counts, mean corpuscular volume, mean corpuscular hemoglobin, and mean corpuscular hemoglobin concentration, as well as dramatically increased reticulocyte counts, serum bilirubin concentrations, and red cell distribution widths. Tissue and organ damage typical of [beta]-thalassemia, such as bone deformities and splenic enlargement due to increased hematopoiesis, are also seen in the heterozygous animals, as is spontaneous iron overload in the spleen, liver, and kidneys. The mice homozygous for the b1 and b2 deletions should be of great value in developing therapies for the treatment of thalassemias in utero. The heterozygous animals will be useful for studying the pathophysiology of thalassemias and have the potential of generating a model of sickle cell anemia when mated with appropriate transgenic animals.

An in vitro transcription assay system, used to study the mechanistic characteristics of transcription activation of the human terminal deoxynucleotide transferase (TdT) indicates that the TATA binding protein enhances the activity of human basal promoter to activate transcription. The primary and secondary basal promoter elements contributed 80% and 20% of the total promoter function. Deletion of the two elements and mutations in the GGGTG sequence present in the transcription start site results in the termination of transcription indicating the regulatory role of the basal promoter elements.

The purpose of crowd counting is to estimate the number of pedestrians in crowd images. Crowd counting or density estimation is an extremely challenging task in computer vision, due to large scale variations and dense scene. Current methods solve these issues by compounding multi-scale Convolutional Neural Network with different receptive fields. In this paper, a novel end-to-end architecture based on Multi-Scale Adversarial Convolutional Neural Network (MSA-CNN) is proposed to generate crowd density and estimate the amount of crowd. Firstly, a multi-scale network is used to extract the globally relevant features in the crowd image, and then fractionally-strided convolutional layers are designed for up-sampling the output to recover the loss of crucial details caused by the earlier max pooling layers. An adversarial loss is directly employed to shrink the estimated value into the realistic subspace to reduce the blurring effect of density estimation. Joint training is performed in an end-to-end fashion using a combination of Adversarial loss and Euclidean loss. The two losses are integrated via a joint training scheme to improve density estimation performance.We conduct some extensive experiments on available datasets to show the significant improvements and supremacy of the proposed approach over the available state-of-the-art approaches. [ABSTRACT FROM AUTHOR]