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Sample records for ace2 null mice

  1. Defective intestinal amino acid absorption in Ace2 null mice.

    PubMed

    Singer, Dustin; Camargo, Simone M R; Ramadan, Tamara; Schäfer, Matthias; Mariotta, Luca; Herzog, Brigitte; Huggel, Katja; Wolfer, David; Werner, Sabine; Penninger, Josef M; Verrey, François

    2012-09-15

    Mutations in the main intestinal and kidney luminal neutral amino acid transporter B(0)AT1 (Slc6a19) lead to Hartnup disorder, a condition that is characterized by neutral aminoaciduria and in some cases pellagra-like symptoms. These latter symptoms caused by low-niacin are thought to result from defective intestinal absorption of its precursor L-tryptophan. Since Ace2 is necessary for intestinal B(0)AT1 expression, we tested the impact of intestinal B(0)AT1 absence in ace2 null mice. Their weight gain following weaning was decreased, and Na(+)-dependent uptake of B(0)AT1 substrates measured in everted intestinal rings was defective. Additionally, high-affinity Na(+)-dependent transport of L-proline, presumably via SIT1 (Slc6a20), was absent, whereas glucose uptake via SGLT1 (Slc5a1) was not affected. Measurements of small intestine luminal amino acid content following gavage showed that more L-tryptophan than other B(0)AT1 substrates reach the ileum in wild-type mice, which is in line with its known lower apparent affinity. In ace2 null mice, the absorption defect was confirmed by a severalfold increase of L-tryptophan and of other neutral amino acids reaching the ileum lumen. Furthermore, plasma and muscle levels of glycine and L-tryptophan were significantly decreased in ace2 null mice, with other neutral amino acids displaying a similar trend. A low-protein/low-niacin diet challenge led to differential changes in plasma amino acid levels in both wild-type and ace2 null mice, but only in ace2 null mice to a stop in weight gain. Despite the combination of low-niacin with a low-protein diet, plasma niacin concentrations remained normal in ace2 null mice and no pellagra symptoms, such as photosensitive skin rash or ataxia, were observed. In summary, mice lacking Ace2-dependent intestinal amino acid transport display no total niacin deficiency nor clear pellagra symptoms, even under a low-protein and low-niacin diet, despite gross amino acid homeostasis alterations.

  2. Regulation of urinary ACE2 in diabetic mice.

    PubMed

    Wysocki, Jan; Garcia-Halpin, Laura; Ye, Minghao; Maier, Christoph; Sowers, Kurt; Burns, Kevin D; Batlle, Daniel

    2013-08-15

    Angiotensin-converting enzyme-2 (ACE2) enhances the degradation of ANG II and its expression is altered in diabetic kidneys, but the regulation of this enzyme in the urine is unknown. Urinary ACE2 was studied in the db/db model of type 2 diabetes and stretozotocin (STZ)-induced type 1 diabetes during several physiological and pharmacological interventions. ACE2 activity in db/db mice was increased in the serum and to a much greater extent in the urine compared with db/m controls. Neither a specific ANG II blocker, telmisartan, nor an ACE inhibitor, captopril, altered the levels of urinary ACE2 in db/db or db/m control mice. High-salt diet (8%) increased whereas low-salt diet (0.1%) decreased urinary ACE2 activity in the urine of db/db mice. In STZ mice, urinary ACE2 was also increased, and insulin decreased it partly but significantly after several weeks of administration. The increase in urinary ACE2 activity in db/db mice reflected an increase in enzymatically active protein with two bands identified of molecular size at 110 and 75 kDa and was associated with an increase in kidney cortex ACE2 protein at 110 kDa but not at 75 kDa. ACE2 activity was increased in isolated tubular preparations but not in glomeruli from db/db mice. Administration of soluble recombinant ACE2 to db/m and db/db mice resulted in a marked increase in serum ACE2 activity, but no gain in ACE2 activity was detectable in the urine, further demonstrating that urinary ACE2 is of kidney origin. Increased urinary ACE2 was associated with more efficient degradation of exogenous ANG II (10(-9) M) in urine from db/db compared with that from db/m mice. Urinary ACE2 could be a potential biomarker of increased metabolism of ANG II in diabetic kidney disease.

  3. Insulin treatment attenuates renal ADAM17 and ACE2 shedding in diabetic Akita mice.

    PubMed

    Salem, Esam S B; Grobe, Nadja; Elased, Khalid M

    2014-03-15

    Angiotensin-converting enzyme 2 (ACE2) is located in several tissues and is highly expressed in renal proximal tubules, where it degrades the vasoconstrictor angiotensin II (ANG II) to ANG-(1-7). Accumulating evidence supports protective roles of ACE2 in several disease states, including diabetic nephropathy. A disintegrin and metalloprotease (ADAM) 17 is involved in the shedding of several transmembrane proteins, including ACE2. Our previous studies showed increased renal ACE2, ADAM17 expression, and urinary ACE2 in type 2 diabetic mice (Chodavarapu H, Grobe N, Somineni HK, Salem ES, Madhu M, Elased KM. PLoS One 8: e62833, 2013). The aim of the present study was to determine the effect of insulin on ACE2 shedding and ADAM17 in type 1 diabetic Akita mice. Results demonstrate increased renal ACE2 and ADAM17 expression and increased urinary ACE2 fragments (≈70 kDa) and albumin excretion in diabetic Akita mice. Immunostaining revealed colocalization of ACE2 with ADAM17 in renal tubules. Renal proximal tubular cells treated with ADAM17 inhibitor showed reduced ACE2 shedding into the media, confirming ADAM17-mediated shedding of ACE2. Treatment of Akita mice with insulin implants for 20 wk normalized hyperglycemia and decreased urinary ACE2 and albumin excretion. Insulin also normalized renal ACE2 and ADAM17 but had no effect on tissue inhibitor of metalloproteinase 3 (TIMP3) protein expression. There was a positive linear correlation between urinary ACE2 and albuminuria, blood glucose, plasma creatinine, glucagon, and triglycerides. This is the first report showing an association between hyperglycemia, cardiovascular risk factors, and increased shedding of urinary ACE2 in diabetic Akita mice. Urinary ACE2 could be used as a biomarker for diabetic nephropathy and as an index of intrarenal ACE2 status.

  4. CD36/Sirtuin 1 Axis Impairment Contributes to Hepatic Steatosis in ACE2-Deficient Mice

    PubMed Central

    Qadri, Fatimunnisa; Penninger, Josef M.; Santos, Robson Augusto S.; Bader, Michael

    2016-01-01

    Background and Aims. Angiotensin converting enzyme 2 (ACE2) is an important component of the renin-angiotensin system. Since angiotensin peptides have been shown to be involved in hepatic steatosis, we aimed to evaluate the hepatic lipid profile in ACE2-deficient (ACE2−/y) mice. Methods. Male C57BL/6 and ACE2−/y mice were analyzed at the age of 3 and 6 months for alterations in the lipid profiles of plasma, faeces, and liver and for hepatic steatosis. Results. ACE2−/y mice showed lower body weight and white adipose tissue at all ages investigated. Moreover, these mice had lower levels of cholesterol, triglycerides, and nonesterified fatty acids in plasma. Strikingly, ACE2−/y mice showed high deposition of lipids in the liver. Expression of CD36, a protein involved in the uptake of triglycerides in liver, was increased in ACE2−/y mice. Concurrently, these mice exhibited an increase in hepatic oxidative stress, evidenced by increased lipid peroxidation and expression of uncoupling protein 2, and downregulation of sirtuin 1. ACE2−/y mice also showed impairments in glucose metabolism and insulin signaling in the liver. Conclusions. Deletion of ACE2 causes CD36/sirtuin 1 axis impairment and thereby interferes with lipid homeostasis, leading to lipodystrophy and steatosis. PMID:28101297

  5. CD36/Sirtuin 1 Axis Impairment Contributes to Hepatic Steatosis in ACE2-Deficient Mice.

    PubMed

    Nunes-Souza, Valéria; Alenina, Natalia; Qadri, Fatimunnisa; Penninger, Josef M; Santos, Robson Augusto S; Bader, Michael; Rabelo, Luiza A

    2016-01-01

    Background and Aims. Angiotensin converting enzyme 2 (ACE2) is an important component of the renin-angiotensin system. Since angiotensin peptides have been shown to be involved in hepatic steatosis, we aimed to evaluate the hepatic lipid profile in ACE2-deficient (ACE2(-/y)) mice. Methods. Male C57BL/6 and ACE2(-/y) mice were analyzed at the age of 3 and 6 months for alterations in the lipid profiles of plasma, faeces, and liver and for hepatic steatosis. Results. ACE2(-/y) mice showed lower body weight and white adipose tissue at all ages investigated. Moreover, these mice had lower levels of cholesterol, triglycerides, and nonesterified fatty acids in plasma. Strikingly, ACE2(-/y) mice showed high deposition of lipids in the liver. Expression of CD36, a protein involved in the uptake of triglycerides in liver, was increased in ACE2(-/y) mice. Concurrently, these mice exhibited an increase in hepatic oxidative stress, evidenced by increased lipid peroxidation and expression of uncoupling protein 2, and downregulation of sirtuin 1. ACE2(-/y) mice also showed impairments in glucose metabolism and insulin signaling in the liver. Conclusions. Deletion of ACE2 causes CD36/sirtuin 1 axis impairment and thereby interferes with lipid homeostasis, leading to lipodystrophy and steatosis.

  6. ACE2 Inhibits Angiotensin II-Induced Abdominal Aortic Aneurysm in Mice.

    PubMed

    Hao, QingQing; Dong, XueFei; Chen, Xu; Yan, Feng; Wang, Xiaoyu; Shi, Haishui; Dong, Bo

    2017-01-31

    Recent study have demonstrated that ACE2 plays an important role in the pathogenesis of abdominal Aortic Aneurysm (AAA). But, little study was reported about the direct effect of ACE2 overexpression on the aneurysm. In this study, we hypothesize that overexpression of ACE2 may prevent the pathogenesis of aneurysm by decreasing RAS activation. Thirty-nine Mice were assigned to 3 groups randomly (n=13 in each group), ACE2 group, Ad.EGFP group and Control group. After 8-week treatment, abdominal aortas with AAA were obtained for HE staining, VVG, immunohistochemistry and Western blotting. The incidence and severity of AAA, macrophage infiltration and MMP protein expression were all detected. The results showed that ACE2 gene transfer significantly decreased the occurrence of AAA and inhibited AAA formation in ApoE-/- mice by inhibiting inflammatory response and MMP activation, the mechanisms may involve decreased ERK and AngII-NF-kB signaling pathways.

  7. Dynamics of ADAM17-Mediated Shedding of ACE2 Applied to Pancreatic Islets of Male db/db Mice

    PubMed Central

    Pedersen, Kim Brint; Chodavarapu, Harshita; Porretta, Constance; Robinson, Leonie K.

    2015-01-01

    Angiotensin-converting enzyme 2 (ACE2) gene therapy aimed at counteracting pancreatic ACE2 depletion improves glucose regulation in two diabetic mouse models: db/db mice and angiotensin II-infused mice. A disintegrin and metalloproteinase 17 (ADAM17) can cause shedding of ACE2 from the cell membrane. The aim of our studies was to determine whether ADAM17 depletes ACE2 levels in pancreatic islets and β-cells. Dynamics of ADAM17-mediated ACE2 shedding were investigated in 832/13 insulinoma cells. Within a wide range of ACE2 expression levels, including the level observed in mouse pancreatic islets, overexpression of ADAM17 increases shed ACE2 and decreases cellular ACE2 levels. We provide a mathematical description of shed and cellular ACE2 activities as a function of the ADAM17 activity. The effect of ADAM17 on the cellular ACE2 content was relatively modest with an absolute control strength value less than 0.25 and approaching 0 at low ADAM17 activities. Although we found that ADAM17 and ACE2 are both expressed in pancreatic islets, the β-cell is not the major cell type expressing ACE2 in islets. During diabetes progression in 8-, 12-, and 15-week-old db/db mice, ACE2 mRNA and ACE2 activity levels in pancreatic islets were not decreased over time nor significantly decreased compared with nondiabetic db/m mice. Levels of ADAM17 mRNA and ADAM17 activity were also not significantly changed. Inhibiting basal ADAM17 activity in mouse islets failed to affect ACE2 levels. We conclude that whereas ADAM17 has the ability to shed ACE2, ADAM17 does not deplete ACE2 from pancreatic islets in diabetic db/db mice. PMID:26441236

  8. ACE2 deficiency reduces β-cell mass and impairs β-cell proliferation in obese C57BL/6 mice

    PubMed Central

    Shoemaker, Robin; Yiannikouris, Frederique; Thatcher, Sean

    2015-01-01

    Drugs that inhibit the renin-angiotensin system (RAS) decrease the onset of type 2 diabetes (T2D). Pancreatic islets express RAS components, including angiotensin-converting enzyme 2 (ACE2), which cleaves angiotensin II (Ang II) to angiotensin-(1–7) [Ang-(1–7)]. Overexpression of ACE2 in pancreas of diabetic mice improved glucose homeostasis. The purpose of this study was to determine if deficiency of endogenous ACE2 contributes to islet dysfunction and T2D. We hypothesized that ACE2 deficiency potentiates the decline in β-cell function and augments the development of diet-induced T2D. Male Ace2+/y or Ace2−/y mice were fed a low-fat (LF) or high-fat (HF) diet for 1 or 4 mo. A subset of 1-mo HF-fed mice were infused with Sal (Sal), losartan (Los), or Ang-(1–7). At 4 mo, while both genotypes of HF-fed mice developed a similar level of insulin resistance, adaptive hyperinsulinemia was reduced in Ace2−/y vs. Ace2+/y mice. Similarly, in vivo glucose-stimulated insulin secretion (GSIS) was reduced in 1-mo HF-fed Ace2−/y compared with Ace2+/y mice, resulting in augmented hyperglycemia. The average islet area was significantly smaller in both LF- and HF-fed Ace2−/y vs. Ace2+/y mice. Additionally, β-cell mass and proliferation were reduced significantly in HF-fed Ace2−/y vs. Ace2+/y mice. Neither infusion of Los nor Ang-(1–7) was able to correct impaired in vivo GSIS of HF-fed ACE2-deficient mice. These results demonstrate a critical role for endogenous ACE2 in the adaptive β-cell hyperinsulinemic response to HF feeding through regulation of β-cell proliferation and growth. PMID:26389599

  9. Administration of 17β-estradiol to ovariectomized obese female mice reverses obesity-hypertension through an ACE2-dependent mechanism.

    PubMed

    Wang, Yu; Shoemaker, Robin; Thatcher, Sean E; Batifoulier-Yiannikouris, Frederique; English, Victoria L; Cassis, Lisa A

    2015-06-15

    We recently demonstrated that female mice are resistant to the development of obesity-induced hypertension through a sex hormone-dependent mechanism that involved adipose angiotensin-converting enzyme 2 (ACE2). In this study, we hypothesized that provision of 17β-estradiol (E2) to ovariectomized (OVX) high-fat (HF)-fed female hypertensive mice would reverse obesity-hypertension through an ACE2-dependent mechanism. Pilot studies defined dose-dependent effects of E2 in OVX female mice on serum E2 concentrations and uterine weights. An E2 dose of 36 μg/ml restored normal serum E2 concentrations and uterine weights. Therefore, HF-fed OVX female Ace2(+/+) and Ace2(-/-) mice were administered vehicle or E2 (36 μg/ml) for 16 wk. E2 administration significantly decreased body weights of HF-fed OVX female Ace2(+/+) and Ace2(-/-) mice of either genotype. At 15 wk, E2 administration decreased systolic blood pressure (SBP) of OVX HF-fed Ace2(+/+) but not Ace2(-/-) females during the light but not the dark cycle. E2-mediated reductions in SBP in Ace2(+/+) females were associated with significant elevations in adipose ACE2 mRNA abundance and activity and reduced plasma ANG II concentrations. In contrast to females, E2 administration had no effect on any parameter quantified in HF-fed male hypertensive mice. In 3T3-L1 adipocytes, E2 promoted ACE2 mRNA abundance through effects at estrogen receptor-α (ERα) and resulted in ERα-mediated binding at the ACE2 promoter. These results demonstrate that E2 administration to OVX females reduces obesity-induced elevations in SBP (light cycle) through an ACE2-dependent mechanism. Beneficial effects of E2 to decrease blood pressure in OVX obese females may result from stimulation of adipose ACE2.

  10. Alternative Roles of STAT3 and MAPK Signaling Pathways in the MMPs Activation and Progression of Lung Injury Induced by Cigarette Smoke Exposure in ACE2 Knockout Mice

    PubMed Central

    Hung, Yi-Han; Hsieh, Wen-Yeh; Hsieh, Jih-Sheng; Liu, Fon-Chang; Tsai, Chin-Hung; Lu, Li-Che; Huang, Chen-Yi; Wu, Chien-Liang; Lin, Chih-Sheng

    2016-01-01

    Inflammation-mediated abnormalities in the renin-angiotensin system (RAS) and expression of matrix metalloproteinases (MMPs) are implicated in the pathogenesis of lung injury. Angiotensin converting enzyme II (ACE2), an angiotensin converting enzyme (ACE) homologue that displays antagonist effects on ACE/angiotensin II (Ang II) axis, could also play a protective role against lung diseases. However, the relationship between ACE2 and MMPs activation in lung injury is still largely unclear. The purpose of this study is to investigate whether MMPs activity could be affected by ACE2 and which ACE2 derived signaling pathways could be also involved via using a mouse model with lung injury induced by cigarette smoke (CS) exposure for 1 to 3 weeks. Wild-type (WT; C57BL/6) and ACE2 KO mice (ACE2-/-) were utilized to study CS-induced lung injury. Increases in the resting respiratory rate (RRR), pulmonary immunokines, leukocyte infiltration and bronchial hyperplasia were observed in the CS-exposed mice. Compared to WT mice, more serious physiopathological changes were found in ACE2-/- mice in the first week of CS exposure. CS exposure increased pulmonary ACE and ACE2 activities in WT mice, and significantly increased ACE in ACE2-/- mice. Furthermore, the activity of pulmonary MMPs was decreased in CS-exposed WT mice, whereas this activity was increased in ACE2-/- mice. CS exposure increased the pulmonary p-p38, p-JNK and p-ERK1/2 level in all mice. In ACE2-/- mice, a significant increase p-STAT3 signaling was detected; however, no effect was observed on the p-STAT3 level in WT mice. Our results support the hypothesis that ACE2 deficiency influences MMPs activation and STAT3 phosphorylation signaling to promote more pulmonary inflammation in the development of lung injury. PMID:27019629

  11. Angiotensin-converting enzyme (ACE and ACE2) imbalance correlates with the severity of cerulein-induced acute pancreatitis in mice.

    PubMed

    Liu, Ruixia; Qi, Haiyu; Wang, Jing; Wang, Yan; Cui, Lijian; Wen, Yan; Yin, Chenghong

    2014-04-01

    Angiotensin-converting enzyme (ACE) and its effector peptide angiotensin II (Ang II) have been implicated in the pathogenesis of pancreatitis. Angiotensin-converting enzyme 2 (ACE2) degrades Ang II to angiotensin-(1-7) [Ang-(1-7)] and has recently been described to have an antagonistic effect on ACE signalling. However, the specific underlying role of ACE2 in the pathogenesis of severe acute pancreatitis (SAP) is unclear. In the present study, the local imbalance of ACE and ACE2, as well as Ang II and Ang-(1-7) expression, was compared in wild-type (WT) and ACE2 knock-out (KO) or ACE2 transgenic (TG) mice subjected to cerulein-induced SAP. Serum amylase, tumour necrosis factor-α, interleukin (IL)-1β, IL-6 and IL-10 levels and histological morphometry were used to determine the severity of pancreatitis. In WT mice, pancreatic ACE and Ang II and serum Ang II expression increased (P < 0.05), while pancreatic ACE2 and Ang-(1-7) and serum Ang-(1-7) levels were also significantly elevated (P < 0.05) from 2 to 72 h after the onset of SAP. However, the ratio of pancreatic ACE2 to ACE expression was significantly reduced (from 1.46 ± 0.09 to 0.27 ± 0.05, P < 0.001) and paralleled the severity of pancreatitis. The Ace2 KO mice exhibited increased levels of tumour necrosis factor-α, IL-1β, IL-6, multifocal coagulative necrosis and inflammatory infiltrate, and lower levels of serum IL-10 and pancreatic Ang-(1-7) (4.70 ± 2.13 versus 10.87 ± 2.51, P < 0.001) compared with cerulein-treated WT mice at the same time point. Conversely, Ace2 TG mice with normal ACE expression were more resistant to SAP challenge as evidenced by a decreased inflammatory response, attenuated pathological changes and increased survival rates. These data suggest that the ACE2-ACE imbalance plays an important role in the pathogenesis of SAP and that pancreatic ACE2 is an important factor in determining the severity of SAP.

  12. ACE2 Deficiency Worsens Epicardial Adipose Tissue Inflammation and Cardiac Dysfunction in Response to Diet-Induced Obesity

    PubMed Central

    Patel, Vaibhav B.; Mori, Jun; McLean, Brent A.; Basu, Ratnadeep; Das, Subhash K.; Ramprasath, Tharmarajan; Parajuli, Nirmal; Penninger, Josef M.; Grant, Maria B.; Lopaschuk, Gary D.

    2016-01-01

    Obesity is increasing in prevalence and is strongly associated with metabolic and cardiovascular disorders. The renin-angiotensin system (RAS) has emerged as a key pathogenic mechanism for these disorders; angiotensin (Ang)-converting enzyme 2 (ACE2) negatively regulates RAS by metabolizing Ang II into Ang 1-7. We studied the role of ACE2 in obesity-mediated cardiac dysfunction. ACE2 null (ACE2KO) and wild-type (WT) mice were fed a high-fat diet (HFD) or a control diet and studied at 6 months of age. Loss of ACE2 resulted in decreased weight gain but increased glucose intolerance, epicardial adipose tissue (EAT) inflammation, and polarization of macrophages into a proinflammatory phenotype in response to HFD. Similarly, human EAT in patients with obesity and heart failure displayed a proinflammatory macrophage phenotype. Exacerbated EAT inflammation in ACE2KO-HFD mice was associated with decreased myocardial adiponectin, decreased phosphorylation of AMPK, increased cardiac steatosis and lipotoxicity, and myocardial insulin resistance, which worsened heart function. Ang 1-7 (24 µg/kg/h) administered to ACE2KO-HFD mice resulted in ameliorated EAT inflammation and reduced cardiac steatosis and lipotoxicity, resulting in normalization of heart failure. In conclusion, ACE2 plays a novel role in heart disease associated with obesity wherein ACE2 negatively regulates obesity-induced EAT inflammation and cardiac insulin resistance. PMID:26224885

  13. Ocular Phenotype of Fbn2-Null Mice

    PubMed Central

    Shi, Yanrong; Tu, Yidong; Mecham, Robert P.; Bassnett, Steven

    2013-01-01

    Purpose. Fibrillin-2 (Fbn2) is the dominant fibrillin isoform expressed during development of the mouse eye. To test its role in morphogenesis, we examined the ocular phenotype of Fbn2−/− mice. Methods. Ocular morphology was assessed by confocal microscopy using antibodies against microfibril components. Results. Fbn2−/− mice had a high incidence of anterior segment dysgenesis. The iris was the most commonly affected tissue. Complete iridal coloboma was present in 37% of eyes. Dyscoria, corectopia and pseudopolycoria were also common (43% combined incidence). In wild-type (WT) mice, fibrillin-2-rich microfibrils are prominent in the pupillary membrane (PM) during development. In Fbn2-null mice, the absence of Fbn2 was partially compensated for by increased expression of fibrillin-1, although the resulting PM microfibrils were disorganized, compared with WTs. In colobomatous adult Fbn2−/− eyes, the PM failed to regress normally, especially beneath the notched region of the iris. Segments of the ciliary body were hypoplastic, and zonular fibers, although relatively plentiful, were unevenly distributed around the lens equator. In regions where the zonular fibers were particularly disturbed, the synchronous differentiation of the underlying lens fiber cells was affected. Conclusions. Fbn2 has an indispensable role in ocular morphogenesis in mice. The high incidence of iris coloboma in Fbn2-null animals implies a previously unsuspected role in optic fissure closure. The observation that fiber cell differentiation was disturbed in Fbn2−/− mice raises the possibility that the attachment of zonular fibers to the lens surface may help specify the equatorial margin of the lens epithelium. PMID:24130178

  14. Context-specific protection of TGFα null mice from osteoarthritis

    PubMed Central

    Usmani, Shirine E.; Ulici, Veronica; Pest, Michael A.; Hill, Tracy L.; Welch, Ian D.; Beier, Frank

    2016-01-01

    Transforming growth factor alpha (TGFα) is a growth factor involved in osteoarthritis (OA). TGFα induces an OA-like phenotype in articular chondrocytes, by inhibiting matrix synthesis and promoting catabolic factor expression. To better understand TGFα’s potential as a therapeutic target, we employed two in vivo OA models: (1) post-traumatic and (2) aging related OA. Ten-week old and six-month old male Tgfa null mice and their heterozygous (control) littermates underwent destabilization of the medial meniscus (DMM) surgery. Disease progression was assessed histologically using the Osteoarthritis Research Society International (OARSI) scoring system. As well, spontaneous disease progression was analyzed in eighteen-month-old Tgfa null and heterozygous mice. Ten-week old Tgfa null mice were protected from OA progression at both seven and fourteen weeks post-surgery. No protection was seen however in six-month old null mice after DMM surgery, and no differences were observed between genotypes in the aging model. Thus, young Tgfa null mice are protected from OA progression in the DMM model, while older mice are not. In addition, Tgfa null mice are equally susceptible to spontaneous OA development during aging. Thus, TGFα might be a valuable therapeutic target in some post-traumatic forms of OA, however its role in idiopathic disease is less clear. PMID:27457421

  15. Context-specific protection of TGFα null mice from osteoarthritis.

    PubMed

    Usmani, Shirine E; Ulici, Veronica; Pest, Michael A; Hill, Tracy L; Welch, Ian D; Beier, Frank

    2016-07-26

    Transforming growth factor alpha (TGFα) is a growth factor involved in osteoarthritis (OA). TGFα induces an OA-like phenotype in articular chondrocytes, by inhibiting matrix synthesis and promoting catabolic factor expression. To better understand TGFα's potential as a therapeutic target, we employed two in vivo OA models: (1) post-traumatic and (2) aging related OA. Ten-week old and six-month old male Tgfa null mice and their heterozygous (control) littermates underwent destabilization of the medial meniscus (DMM) surgery. Disease progression was assessed histologically using the Osteoarthritis Research Society International (OARSI) scoring system. As well, spontaneous disease progression was analyzed in eighteen-month-old Tgfa null and heterozygous mice. Ten-week old Tgfa null mice were protected from OA progression at both seven and fourteen weeks post-surgery. No protection was seen however in six-month old null mice after DMM surgery, and no differences were observed between genotypes in the aging model. Thus, young Tgfa null mice are protected from OA progression in the DMM model, while older mice are not. In addition, Tgfa null mice are equally susceptible to spontaneous OA development during aging. Thus, TGFα might be a valuable therapeutic target in some post-traumatic forms of OA, however its role in idiopathic disease is less clear.

  16. Angiogenesis in Platelet Endothelial Cell Adhesion Molecule-1-Null Mice

    PubMed Central

    Cao, Gaoyuan; Fehrenbach, Melane L.; Williams, James T.; Finklestein, Jeffrey M.; Zhu, Jing-Xu; DeLisser, Horace M.

    2009-01-01

    Platelet endothelial cell adhesion molecule (PECAM)-1 has been previously implicated in endothelial cell migration; additionally, anti-PECAM-1 antibodies have been shown to inhibit in vivo angiogenesis. Studies were therefore performed with PECAM-1-null mice to further define the involvement of PECAM-1 in blood vessel formation. Vascularization of subcutaneous Matrigel implants as well as tumor angiogenesis were both inhibited in PECAM-1-null mice. Reciprocal bone marrow transplants that involved both wild-type and PECAM-1-deficient mice revealed that the impaired angiogenic response resulted from a loss of endothelial, but not leukocyte, PECAM-1. In vitro wound migration and single-cell motility by PECAM-1-null endothelial cells were also compromised. In addition, filopodia formation, a feature of motile cells, was inhibited in PECAM-1-null endothelial cells as well as in human endothelial cells treated with either anti-PECAM-1 antibody or PECAM-1 siRNA. Furthermore, the expression of PECAM-1 promoted filopodia formation and increased the protein expression levels of Cdc42, a Rho GTPase that is known to promote the formation of filopodia. In the developing retinal vasculature, numerous, long filamentous filopodia, emanating from endothelial cells at the tips of angiogenic sprouts, were observed in wild-type animals, but to a lesser extent in the PECAM-1-null mice. Together, these data further establish the involvement of endothelial PECAM-1 in angiogenesis and suggest that, in vivo, PECAM-1 may stimulate endothelial cell motility by promoting the formation of filopodia. PMID:19574426

  17. sirt1-null mice develop an autoimmune-like condition

    SciTech Connect

    Sequeira, Jedon; Boily, Gino; Bazinet, Stephanie; Saliba, Sarah; He Xiaohong; Jardine, Karen; Kennedy, Christopher; Staines, William; Rousseaux, Colin; Mueller, Rudi; McBurney, Michael W.

    2008-10-01

    The sirt1 gene encodes a protein deacetylase with a broad spectrum of reported substrates. Mice carrying null alleles for sirt1 are viable on outbred genetic backgrounds so we have examined them in detail to identify the biological processes that are dependent on SIRT1. Sera from adult sirt1-null mice contain antibodies that react with nuclear antigens and immune complexes become deposited in the livers and kidneys of these animals. Some of the sirt1-null animals develop a disease resembling diabetes insipidus when they approach 2 years of age although the relationship to the autoimmunity remains unclear. We interpret these observations as consistent with a role for SIRT1 in sustaining normal immune function and in this way delaying the onset of autoimmune disease.

  18. Brain ACE2 shedding contributes to the development of neurogenic hypertension

    PubMed Central

    Chhabra, Kavaljit H.; Lazartigues, Eric

    2015-01-01

    Rationale Over-activity of the brain Renin Angiotensin System (RAS) is a major contributor to neurogenic hypertension. While over-expression of Angiotensin-Converting Enzyme type 2 (ACE2) has been shown to be beneficial in reducing hypertension by transforming Angiotensin (Ang)-II into Ang-(1-7), several groups have reported decreased brain ACE2 expression and activity during the development of hypertension. Objective We hypothesized that ADAM17-mediated ACE2 shedding results in decreased membrane-bound ACE2 in the brain, thus promoting the development of neurogenic hypertension. Methods and Results To test this hypothesis, we used the DOCA-salt model of neurogenic hypertension in non-transgenic (NT) and syn-hACE2 mice over-expressing ACE2 in neurons. DOCA-salt treatment in NT mice led to significant increases in blood pressure, hypothalamic Ang-II levels, inflammation, impaired baroreflex sensitivity, autonomic dysfunction, as well as decreased hypothalamic ACE2 activity and expression, while these changes were blunted or prevented in syn-hACE2 mice. In addition, reduction of ACE2 expression and activity in the brain paralleled a rise in ACE2 activity in the cerebrospinal fluid of NT mice following DOCA-salt treatment and was accompanied by enhanced ADAM17 expression and activity in the hypothalamus. Chronic knockdown of ADAM17 in the brain blunted the development of hypertension and restored ACE2 activity and baroreflex function. Conclusions Our data provide the first evidence that ADAM17-mediated shedding impairs brain ACE2 compensatory activity, thus contributing to the development of neurogenic hypertension. PMID:24014829

  19. The Aspergillus fumigatus Transcription Factor Ace2 Governs Pigment Production, Conidiation and Virulence

    PubMed Central

    Ejzykowicz, Daniele E.; Cunha, Marcel M.; Rozental, Sonia; Solis, Norma V.; Gravelat, Fabrice N.; Sheppard, Donald C.; Filler, Scott G.

    2009-01-01

    Summary Aspergillus fumigatus causes serious and frequently fatal infections in immunocompromised patients. To investigate the regulation of virulence of this fungus, we constructed and analyzed an A. fumigatus mutant that lacked the transcription factor Ace2, which influences virulence in other fungi. The Δace2 mutant had dysmorphic conidiophores, reduced conidia production, and abnormal conidial cell wall architecture. This mutant produced an orange pigment when grown on solid media, although its conidia had normal pigmentation. Conidia of the Δace2 mutant were larger and had accelerated germination. The resulting germlings were resistant to hydrogen peroxide, but not other stressors. Non-neutropenic mice that were immunosuppressed with cortisone acetate and infected with the Δace2 mutant had accelerated mortality, greater pulmonary fungal burden, and increased pulmonary inflammatory responses compared to mice infected with the wild-type or Δace2ace2 complemented strains. The Δace2 mutant had reduced ppoC, ecm33, and ags3 mRNA expression. It is known that A. fumigatus mutants with absent or reduced expression of these genes have increased virulence in mice, as well as other phenotypic similarities to the Δace2 mutant. Therefore, reduced expression of these genes likely contributes to the increased virulence of the Δace2 mutant. PMID:19220748

  20. Cadmium modulates adipocyte functions in metallothionein-null mice

    SciTech Connect

    Kawakami, Takashige; Nishiyama, Kaori; Kadota, Yoshito; Sato, Masao; Inoue, Masahisa; Suzuki, Shinya

    2013-11-01

    Our previous study has demonstrated that exposure to cadmium (Cd), a toxic heavy metal, causes a reduction of adipocyte size and the modulation of adipokine expression. To further investigate the significance of the Cd action, we studied the effect of Cd on the white adipose tissue (WAT) of metallothionein null (MT{sup −/−}) mice, which cannot form atoxic Cd–MT complexes and are used for evaluating Cd as free ions, and wild type (MT{sup +/+}) mice. Cd administration more significantly reduced the adipocyte size of MT{sup −/−} mice than that of MT{sup +/+} mice. Cd exposure also induced macrophage recruitment to WAT with an increase in the expression level of Ccl2 (MCP-1) in the MT{sup −/−} mice. The in vitro exposure of Cd to adipocytes induce triglyceride release into culture medium, decrease in the expression levels of genes involved in fatty acid synthesis and lipid hydrolysis at 24 h, and at 48 h increase in phosphorylation of the lipid-droplet-associated protein perilipin, which facilitates the degradation of stored lipids in adipocytes. Therefore, the reduction in adipocyte size by Cd may arise from an imbalance between lipid synthesis and lipolysis. In addition, the expression levels of leptin, adiponectin and resistin decreased in adipocytes. Taken together, exposure to Cd may induce unusually small adipocytes and modulate the expression of adipokines differently from the case of physiologically small adipocytes, and may accelerate the risk of developing insulin resistance and type 2 diabetes. - Highlights: • Cd causes a marked reduction in adipocyte size in MT-null mice. • Cd enhances macrophage migration into adipose tissue and disrupt adipokine secretion. • MT gene alleviates Cd-induced adipocyte dysfunctions. • Cd enhances the degradation of stored lipids in adipocytes, mediated by perilipin. • Cd induces unusually small adipocytes and the abnormal expression of adipokines.

  1. Mechanical Forces Exacerbate Periodontal Defects in Bsp-null Mice

    PubMed Central

    Soenjaya, Y.; Foster, B.L.; Nociti, F.H.; Ao, M.; Holdsworth, D.W.; Hunter, G.K.; Somerman, M.J.

    2015-01-01

    Bone sialoprotein (BSP) is an acidic phosphoprotein with collagen-binding, cell attachment, and hydroxyapatite-nucleating properties. BSP expression in mineralized tissues is upregulated at onset of mineralization. Bsp-null (Bsp-/-) mice exhibit reductions in bone mineral density, bone turnover, osteoclast activation, and impaired bone healing. Furthermore, Bsp-/- mice have marked periodontal tissue breakdown, with a lack of acellular cementum leading to periodontal ligament detachment, extensive alveolar bone and tooth root resorption, and incisor malocclusion. We hypothesized that altered mechanical stress from mastication contributes to periodontal destruction observed in Bsp-/- mice. This hypothesis was tested by comparing Bsp-/- and wild-type mice fed with standard hard pellet diet or soft powder diet. Dentoalveolar tissues were analyzed using histology and micro–computed tomography. By 8 wk of age, Bsp-/- mice exhibited molar and incisor malocclusion regardless of diet. Bsp-/- mice with hard pellet diet exhibited high incidence (30%) of severe incisor malocclusion, 10% lower body weight, 3% reduced femur length, and 30% elevated serum alkaline phosphatase activity compared to wild type. Soft powder diet reduced severe incisor malocclusion incidence to 3% in Bsp-/- mice, supporting the hypothesis that occlusal loading contributed to the malocclusion phenotype. Furthermore, Bsp-/- mice in the soft powder diet group featured normal body weight, long bone length, and serum alkaline phosphatase activity, suggesting that tooth dysfunction and malnutrition contribute to growth and skeletal defects reported in Bsp-/- mice. Bsp-/- incisors also erupt at a slower rate, which likely leads to the observed thickened dentin and enhanced mineralization of dentin and enamel toward the apical end. We propose that the decrease in eruption rate is due to a lack of acellular cementum and associated defective periodontal attachment. These data demonstrate the importance of BSP

  2. Mechanical Forces Exacerbate Periodontal Defects in Bsp-null Mice.

    PubMed

    Soenjaya, Y; Foster, B L; Nociti, F H; Ao, M; Holdsworth, D W; Hunter, G K; Somerman, M J; Goldberg, H A

    2015-09-01

    Bone sialoprotein (BSP) is an acidic phosphoprotein with collagen-binding, cell attachment, and hydroxyapatite-nucleating properties. BSP expression in mineralized tissues is upregulated at onset of mineralization. Bsp-null (Bsp(-/-)) mice exhibit reductions in bone mineral density, bone turnover, osteoclast activation, and impaired bone healing. Furthermore, Bsp(-/-) mice have marked periodontal tissue breakdown, with a lack of acellular cementum leading to periodontal ligament detachment, extensive alveolar bone and tooth root resorption, and incisor malocclusion. We hypothesized that altered mechanical stress from mastication contributes to periodontal destruction observed in Bsp(-/-) mice. This hypothesis was tested by comparing Bsp(-/-) and wild-type mice fed with standard hard pellet diet or soft powder diet. Dentoalveolar tissues were analyzed using histology and micro-computed tomography. By 8 wk of age, Bsp(-/-) mice exhibited molar and incisor malocclusion regardless of diet. Bsp(-/-) mice with hard pellet diet exhibited high incidence (30%) of severe incisor malocclusion, 10% lower body weight, 3% reduced femur length, and 30% elevated serum alkaline phosphatase activity compared to wild type. Soft powder diet reduced severe incisor malocclusion incidence to 3% in Bsp(-/-) mice, supporting the hypothesis that occlusal loading contributed to the malocclusion phenotype. Furthermore, Bsp(-/-) mice in the soft powder diet group featured normal body weight, long bone length, and serum alkaline phosphatase activity, suggesting that tooth dysfunction and malnutrition contribute to growth and skeletal defects reported in Bsp(-/-) mice. Bsp(-/-) incisors also erupt at a slower rate, which likely leads to the observed thickened dentin and enhanced mineralization of dentin and enamel toward the apical end. We propose that the decrease in eruption rate is due to a lack of acellular cementum and associated defective periodontal attachment. These data demonstrate the

  3. ACE2 alterations in kidney disease

    PubMed Central

    Soler, María José; Wysocki, Jan; Batlle, Daniel

    2013-01-01

    Angiotensin-converting enzyme 2 (ACE2) is a monocarboxypeptidase that degrades angiotensin (Ang) II to Ang-(1–7). ACE2 is highly expressed within the kidneys, it is largely localized in tubular epithelial cells and less prominently in glomerular epithelial cells and in the renal vasculature. ACE2 activity has been shown to be altered in diabetic kidney disease, hypertensive renal disease and in different models of kidney injury. There is often a dissociation between tubular and glomerular ACE2 expression, particularly in diabetic kidney disease where ACE2 expression is increased at the tubular level but decreased at the glomerular level. In this review, we will discuss alterations in circulating and renal ACE2 recently described in different renal pathologies and disease models as well as their possible significance. PMID:23956234

  4. Neuropathy in Human and Mice with PMP22 null

    PubMed Central

    Saporta, Mario Andre; Katona, Istvan; Zhang, Xuebao; Roper, Helen P.; Carr, Louise; Macdonald, Fiona; Brueton, Louise; Blake, Julian; Suter, Ueli; Reilly, Mary M.; Shy, Michael E.; Li, Jun

    2013-01-01

    Background/Objective Haploinsufficiency of PMP22 causes hereditary neuropathy with liability to pressure palsies (HNPP). However, the biological functions of PMP22 in humans are largely unexplored due to the absence of patients with PMP22 null mutations. Design, Setting and Participants We have evaluated a 7-year-old boy with PMP22 null. Findings were compared with those from nerves of Pmp22 null mice. Results Motor and sensory deficits in the proband were non-length dependent. Weakness was found in cranial muscles, but not in the limbs. Large fiber sensory modalities were profoundly abnormal, which started prior to the maturation of myelin. This is in line with the temporal pattern of PMP22 expression predominantly in cranial motor neurons and DRG during embryonic development, becoming undetectable in adulthood. Moreover, there were conspicuous maturation defects of myelinating Schwann cells that were more significant in motor nerve fibers than in sensory nerve fibers. Conclusions Taken together, these data suggest that PMP22 is important for the normal function of neurons that express PMP22 during early development, such as cranial motor neurons and spinal sensory neurons. Moreover, PMP22 deficiency differentially affects myelination between motor and sensory nerves, which may have contributed to the unique clinical phenotype in the patient with absence of PMP22. PMID:21670407

  5. ACE2 is augmented in dystrophic skeletal muscle and plays a role in decreasing associated fibrosis.

    PubMed

    Riquelme, Cecilia; Acuña, María José; Torrejón, Javiera; Rebolledo, Daniela; Cabrera, Daniel; Santos, Robson A; Brandan, Enrique

    2014-01-01

    Duchenne muscular dystrophy (DMD) is the most common inherited neuromuscular disease and is characterized by absence of the cytoskeletal protein dystrophin, muscle wasting, and fibrosis. We previously demonstrated that systemic infusion or oral administration of angiotensin-(1-7) (Ang-(1-7)), a peptide with opposing effects to angiotensin II, normalized skeletal muscle architecture, decreased local fibrosis, and improved muscle function in mdx mice, a dystrophic model for DMD. In this study, we investigated the presence, activity, and localization of ACE2, the enzyme responsible for Ang-(1-7) production, in wild type (wt) and mdx skeletal muscle and in a model of induced chronic damage in wt mice. All dystrophic muscles studied showed higher ACE2 activity than wt muscle. Immunolocalization studies indicated that ACE2 was localized mainly at the sarcolemma and, to a lesser extent, associated with interstitial cells. Similar results were observed in the model of chronic damage in the tibialis anterior (TA) muscle. Furthermore, we evaluated the effect of ACE2 overexpression in mdx TA muscle using an adenovirus containing human ACE2 sequence and showed that expression of ACE2 reduced the fibrosis associated with TA dystrophic muscles. Moreover, we observed fewer inflammatory cells infiltrating the mdx muscle. Finally, mdx gastrocnemius muscles from mice infused with Ang-(1-7), which decreases fibrosis, contain less ACE2 associated with the muscle. This is the first evidence supporting ACE2 as an important therapeutic target to improve the dystrophic skeletal muscle phenotype.

  6. Development of pheochromocytoma in ceramide synthase 2 null mice.

    PubMed

    Park, Woo-Jae; Brenner, Ori; Kogot-Levin, Aviram; Saada, Ann; Merrill, Alfred H; Pewzner-Jung, Yael; Futerman, Anthony H

    2015-08-01

    Pheochromocytoma (PCC) and paraganglioma are rare neuroendocrine tumors of the adrenal medulla and sympathetic and parasympathetic paraganglia, for which mutations in ∼15 disease-associated genes have been identified. We now document the role of an additional gene in mice, the ceramide synthase 2 (CerS2) gene. CerS2, one of six mammalian CerS, synthesizes ceramides with very-long (C22-C24) chains. The CerS2 null mouse has been well characterized and displays lesions in several organs including the liver, lung and the brain. We now demonstrate that changes in the sphingolipid acyl chain profile of the adrenal gland lead to the generation of adrenal medullary tumors. Histological analyses revealed that about half of the CerS2 null mice developed PCC by ∼13 months, and the rest showed signs of medullary hyperplasia. Norepinephrine and normetanephrine levels in the urine were elevated at 7 months of age consistent with the morphological abnormalities found at later ages. Accumulation of ceroid in the X-zone was observed as early as 2 months of age and as a consequence, older mice displayed elevated levels of lysosomal cathepsins, reduced proteasome activity and reduced activity of mitochondrial complex IV by 6 months of age. Together, these findings implicate an additional pathway that can lead to PCC formation, which involves alterations in the sphingolipid acyl chain length. Analysis of the role of sphingolipids in PCC may lead to further understanding of the mechanism by which PCC develops, and might implicate the sphingolipid pathway as a possible novel therapeutic target for this rare tumor.

  7. Renal NHE expression and activity in neonatal NHE3- and NHE8-null mice.

    PubMed

    Pirojsakul, Kwanchai; Gattineni, Jyothsna; Dwarakanath, Vangipuram; Baum, Michel

    2015-01-01

    Na(+)/H(+) exchanger (NHE)3 is the predominant NHE on the brush-border membrane of the proximal tubule in adult animals. NHE8 has been localized to the brush-border membrane of proximal tubules and is more highly expressed in neonates than in adult animals. However, the relative role of NHE8 in neonatal renal acidification is unclear. The present study examined if there was a compensatory increase in NHE3 in NHE8-null neonatal mice and whether there was a compensatory increase in NHE8 in NHE3-null neonatal mice. In addition, we examined whether wild-type, NHE3-null, and NHE8-null mice had an increase in NHE activity in response to metabolic acidosis. We found that at baseline, there was comparable renal NHE3 mRNA, total protein, and brush-border membrane protein abundance as in neonatal control and NHE8-null mice. There was comparable renal NHE8 mRNA, total protein, and brush-border membrane protein abundance in NHE3-null neonatal and control mice. Both NHE3- and NHE8-null mice had a comparable but lower rate of NHE activity than control mice. We next imposed metabolic acidosis in wild-type, NHE3-null, and NHE8-null mice. Acidemic NHE8-null mice had an increase in brush-border membrane vesicle NHE3 protein abundance and NHE activity compared with vehicle-treated mice. Likewise, NHE3-null mice had an increase in NHE8 brush-border membrane protein abundance and NHE activity in response to metabolic acidosis. In conclusion, both NHE3 and NHE8 likely play a role in neonatal acidification.

  8. Interaction of diabetes and ACE2 in the pathogenesis of cardiovascular disease in experimental diabetes.

    PubMed

    Tikellis, Chris; Pickering, Raelene; Tsorotes, Despina; Du, Xiao-Jun; Kiriazis, Helen; Nguyen-Huu, Thu-Phuc; Head, Geoffrey A; Cooper, Mark E; Thomas, Merlin C

    2012-10-01

    Local and systemic AngII (angiotensin II) levels are regulated by ACE2 (angiotensin-converting enzyme 2), which is reduced in diabetic tissues. In the present study, we examine the effect of ACE2 deficiency on the early cardiac and vascular changes associated with experimental diabetes. Streptozotocin diabetes was induced in male C57BL6 mice and Ace2-KO (knockout) mice, and markers of RAS (renin-angiotensin system) activity, cardiac function and injury were assessed after 10 weeks. In a second protocol, diabetes was induced in male ApoE (apolipoprotein E)-KO mice and ApoE/Ace2-double-KO mice, and plaque accumulation and markers of atherogenesis assessed after 20 weeks. The induction of diabetes in wild-type mice led to reduced ACE2 expression and activity in the heart, elevated circulating AngII levels and reduced cardiac Ang-(1-7) [angiotensin-(1-7)] levels. This was associated structurally with thinning of the LV (left ventricular) wall and mild ventricular dilatation, and histologically with increased cardiomyocyte apoptosis on TUNEL (terminal deoxynucleotidyl transferase-mediated dUTP nick-end labelling) staining and compensatory hypertrophy denoted by an increased cardiomyocyte cross-sectional area. By contrast Ace2-KO mice failed to increase circulating AngII concentration, experienced a paradoxical fall in cardiac AngII levels and no change in Ang-(1-7) following the onset of diabetes. At the same time the major phenotypic differences between Ace2-deficient and Ace2-replete mice with respect to BP (blood pressure) and cardiac hypertrophy were eliminated following the induction of diabetes. Consistent with findings in the heart, the accelerated atherosclerosis that was observed in diabetic ApoE-KO mice was not seen in diabetic ApoE/Ace2-KO mice, which experienced no further increase in plaque accumulation or expression in key adhesion molecules beyond that seen in ApoE/Ace2-KO mice. These results point to the potential role of ACE2 deficiency in regulating

  9. Connexin43 null mice reveal that astrocytes express multiple connexins.

    PubMed

    Dermietzel, R; Gao, Y; Scemes, E; Vieira, D; Urban, M; Kremer, M; Bennett, M V; Spray, D C

    2000-04-01

    The gap junction protein connexin43 (Cx43) is the primary component of intercellular channels in cardiac tissue and in astrocytes, the most abundant type of glial cells in the brain. Mice in which the gene for Cx43 is deleted by homologous recombination die at birth, due to profound hypertrophy of the ventricular outflow tract and stenosis of the pulmonary artery. Despite this significant cardiovascular abnormality, brains of connexin43 null [Cx43 (-/-)] animals are shown to be macroscopically normal and to display a pattern of cortical lamination that is not detectably different from wildtype siblings. Presence of Cx40 and Cx45 in brains and astrocytes cultured from both Cx43 (-/-) mice and wildtype littermates was confirmed by RT-PCR, Northern blot analyses and by immunostaining; Cx46 was detected by RT-PCR and Northern blot analyses. Presence of Cx26 in astrocyte cultures was indicated by RT-PCR and by Western blot analysis, although we were unable to resolve whether it was contributed by contaminating cells; Cx30 mRNA was detected by Northern blot in long term (2 weeks) but not fresh cultures of astrocytes. These studies thus reveal that astrocyte gap junctions may be formed of multiple connexins. Presumably, the metabolic and ionic coupling provided by these diverse gap junction types may functionally compensate for the absence of the major astrocyte gap junction protein in Cx43 (-/-) mice, providing whatever intercellular signaling is necessary for brain development and cortical lamination.

  10. Muscular dystrophy in PTFR/cavin-1 null mice

    PubMed Central

    Ding, Shi-Ying; Pilch, Paul F.

    2017-01-01

    ice and humans lacking the caveolae component polymerase I transcription release factor (PTRF, also known as cavin-1) exhibit lipo- and muscular dystrophy. Here we describe the molecular features underlying the muscle phenotype for PTRF/cavin-1 null mice. These animals had a decreased ability to exercise, and exhibited muscle hypertrophy with increased muscle fiber size and muscle mass due, in part, to constitutive activation of the Akt pathway. Their muscles were fibrotic and exhibited impaired membrane integrity accompanied by an apparent compensatory activation of the dystrophin-glycoprotein complex along with elevated expression of proteins involved in muscle repair function. Ptrf deletion also caused decreased mitochondrial function, oxygen consumption, and altered myofiber composition. Thus, in addition to compromised adipocyte-related physiology, the absence of PTRF/cavin-1 in mice caused a unique form of muscular dystrophy with a phenotype similar or identical to that seen in humans lacking this protein. Further understanding of this muscular dystrophy model will provide information relevant to the human situation and guidance for potential therapies. PMID:28289716

  11. Reduced arsenic clearance and increased toxicity in aquaglyceroporin-9-null mice

    PubMed Central

    Carbrey, Jennifer M.; Song, Linhua; Zhou, Yao; Yoshinaga, Masafumi; Rojek, Aleksandra; Wang, Yiding; Liu, Yangjian; Lujan, Heidi L.; DiCarlo, Stephen E.; Nielsen, Søren; Rosen, Barry P.; Agre, Peter; Mukhopadhyay, Rita

    2009-01-01

    Expressed in liver, aquaglyceroporin-9 (AQP9) is permeated by glycerol, arsenite, and other small, neutral solutes. To evaluate a possible protective role, AQP9-null mice were evaluated for in vivo arsenic toxicity. After injection with NaAsO2, AQP9-null mice suffer reduced survival rates (LD50, 12 mg/kg) compared with WT mice (LD50, 15 mg/kg). The highest tissue level of arsenic is in heart, with AQP9-null mice accumulating 10–20 times more arsenic than WT mice. Within hours after NaAsO2 injection, AQP9-null mice sustain profound bradycardia, despite normal serum electrolytes. Increased arsenic levels are also present in liver, lung, spleen, and testis of AQP9-null mice. Arsenic levels in the feces and urine of AQP9-null mice are only ≈10% of the WT levels, and reduced clearance of multiple arsenic species by the AQP9-null mice suggests that AQP9 is involved in the export of multiple forms of arsenic. Immunohistochemical staining of liver sections revealed that AQP9 is most abundant in basolateral membrane of hepatocytes adjacent to the sinusoids. AQP9 is not detected in heart or kidney by PCR or immunohistochemistry. We propose that AQP9 provides a route for excretion of arsenic by the liver, thereby providing partial protection of the whole animal from arsenic toxicity. PMID:19805235

  12. ACE and ACE2 in kidney disease

    PubMed Central

    Mizuiri, Sonoo; Ohashi, Yasushi

    2015-01-01

    Renin angiotensin system (RAS) activation has a significant influence on renal disease progression. The classical angiotensin-converting enzyme (ACE)-angiotensin II (Ang II)-Ang II type 1 (AT1) axis is considered to control the effects of RAS activation on renal disease. However, since its discovery in 2000 ACE2 has also been demonstrated to have a significant impact on the RAS. The synthesis and catabolism of Ang II are regulated via a complex series of interactions, which involve ACE and ACE2. In the kidneys, ACE2 is expressed in the proximal tubules and less strongly in the glomeruli. The synthesis of inactive Ang 1-9 from Ang I and the catabolism of Ang II to produce Ang 1-7 are the main functions of ACE2. Ang 1-7 reduces vasoconstriction, water retention, salt intake, cell proliferation, and reactive oxygen stress, and also has a renoprotective effect. Thus, in the non-classical RAS the ACE2-Ang 1-7-Mas axis counteracts the ACE-Ang II-AT1 axis. This review examines recent human and animal studies about renal ACE and ACE2. PMID:25664248

  13. Angiotensin-II mediates ACE2 Internalization and Degradation through an Angiotensin-II type I receptor-dependent mechanism

    PubMed Central

    Lazartigues, Eric; Filipeanu, Catalin M.

    2014-01-01

    Angiotensin Converting Enzyme type 2 (ACE2) is a pivotal component of the renin-angiotensin system, promoting the conversion of Angiotensin (Ang)-II to Ang-(1-7). We previously reported that decreased ACE2 expression and activity contribute to the development of Ang-II-mediated hypertension in mice. The present study aimed to investigate the mechanisms involved in ACE2 down-regulation during neurogenic hypertension. In ACE2-transfected Neuro-2A cells, Ang-II treatment resulted in a significant attenuation of ACE2 enzymatic activity. Examination of the subcellular localization of ACE2 revealed that Ang-II treatment leads to ACE2 internalization and degradation into lysosomes. These effects were prevented by both the Ang-II type 1 receptor (AT1R) blocker losartan and the lysosomal inhibitor leupeptin. In contrast, in HEK293T cells, which lack endogenous AT1R, Ang-II failed to promote ACE2 internalization. Moreover, this effect could be induced after AT1R transfection. Further, co-immunoprecipitation experiments demonstrated that AT1R and ACE2 form complexes and these interactions were decreased by Ang-II treatment, which also enhanced ACE2 ubiquitination. In contrast, ACE2 activity was not changed by transfection of AT2 or Mas receptors. In vivo, Ang-II-mediated hypertension was blunted by chronic infusion of leupeptin in wildtype C57Bl/6, but not in ACE2 knockout mice. Overall, this is the first demonstration that elevated Ang-II levels reduce ACE2 expression and activity by stimulation of lysosomal degradation through an AT1R-dependent mechanism. PMID:25225202

  14. Characterization of ACE and ACE2 Expression within Different Organs of the NOD Mouse.

    PubMed

    Roca-Ho, Heleia; Riera, Marta; Palau, Vanesa; Pascual, Julio; Soler, Maria Jose

    2017-03-05

    Renin angiotensin system (RAS) is known to play a key role in several diseases such as diabetes, and renal and cardiovascular pathologies. Its blockade has been demonstrated to delay chronic kidney disease progression and cardiovascular damage in diabetic patients. In this sense, since local RAS has been described, the aim of this study is to characterize angiotensin converting enzyme (ACE) and ACE2 activities, as well as protein expression, in several tissues of the non-obese diabetic (NOD) mice model. After 21 or 40 days of diabetes onset, mouse serums and tissues were analyzed for ACE and ACE2 enzyme activities and protein expression. ACE and ACE2 enzyme activities were detected in different tissues. Their expressions vary depending on the studied tissue. Thus, whereas ACE activity was highly expressed in lungs, ACE2 activity was highly expressed in pancreas among the studied tissues. Interestingly, we also observed that diabetes up-regulates ACE mainly in serum, lung, heart, and liver, and ACE2 mainly in serum, liver, and pancreas. In conclusion, we found a marked serum and pulmonary alteration in ACE activity of diabetic mice, suggesting a common regulation. The increase of ACE2 activity within the circulation in diabetic mice may be ascribed to a compensatory mechanism of RAS.

  15. Characterization of ACE and ACE2 Expression within Different Organs of the NOD Mouse

    PubMed Central

    Roca-Ho, Heleia; Riera, Marta; Palau, Vanesa; Pascual, Julio; Soler, Maria Jose

    2017-01-01

    Renin angiotensin system (RAS) is known to play a key role in several diseases such as diabetes, and renal and cardiovascular pathologies. Its blockade has been demonstrated to delay chronic kidney disease progression and cardiovascular damage in diabetic patients. In this sense, since local RAS has been described, the aim of this study is to characterize angiotensin converting enzyme (ACE) and ACE2 activities, as well as protein expression, in several tissues of the non-obese diabetic (NOD) mice model. After 21 or 40 days of diabetes onset, mouse serums and tissues were analyzed for ACE and ACE2 enzyme activities and protein expression. ACE and ACE2 enzyme activities were detected in different tissues. Their expressions vary depending on the studied tissue. Thus, whereas ACE activity was highly expressed in lungs, ACE2 activity was highly expressed in pancreas among the studied tissues. Interestingly, we also observed that diabetes up-regulates ACE mainly in serum, lung, heart, and liver, and ACE2 mainly in serum, liver, and pancreas. In conclusion, we found a marked serum and pulmonary alteration in ACE activity of diabetic mice, suggesting a common regulation. The increase of ACE2 activity within the circulation in diabetic mice may be ascribed to a compensatory mechanism of RAS. PMID:28273875

  16. Cardiac Characterization of sgca-Null Mice Using High Resolution Echocardiography.

    PubMed

    Fayssoil, Abdallah; Renault, Gilles; Guerchet, Nicolas; Marchiol-Fournigault, Carmen; Fougerousse, Françoise; Richard, Isabelle

    2013-01-01

    Limb-girdle muscular dystrophy 2D (LGMD2D) is an inherited myogenic disorder belonging to the group of muscular dystrophies. Sgca-null mouse is a knock-out model of LGMD2D. Little is known about cardiac phenotype characterization in this model at different ages. We conducted a prospective study to characterize cardiac sgca-null mice phenotype using high resolution Doppler echocardiography at different ages. Conventional echocardiography was performed on anesthetised mice using a Vevo 770 (Visualsonics) with 30 MHz cardiac probe. Wild Type (WT) and sgca-null mice were scanned at 13, 15 and 17 months. From M-mode, we measured interventricular septal (IVS) wall thickness, posterior wall (PW) thickness, and end-left ventricular diameter in systolic and diastolic. From the above parameters, we calculated left ventricular (LV) shortening fraction (SF), LV ejection fraction (EF) and LV mass. At age 13 months, PW diastolic thickness was increased in sgca-null mice (0.89±0.14 mm vs 0.73±0.2 mm; P=0.020) and LV mass was higher in sgca-null mice (LV mass 205.2 mg vs 143 mg; P=0.001). We found also dilation of the LV (LVEDD: 4.84 mm vs 4.29 mm; P=0.019) in sgca-null mice. At age 15 months, dilation of the LV (LVEDD: 4.86 mm vs 4 mm; P=0.05) with an increase of the LV mass (165.7 mg vs 127.12; P=0.03) are found in sgca-null mice. At age 17 months, we found a decrease of the PW thickening (17% vs 30%; P=0.036). This work provides echocardiographic insights for the assessment of pharmaceutical therapies in sgca-null mice.

  17. Cardiac Characterization of sgca-Null Mice Using High Resolution Echocardiography

    PubMed Central

    Fayssoil, Abdallah; Renault, Gilles; Guerchet, Nicolas; Marchiol-Fournigault, Carmen; Fougerousse, Françoise; Richard, Isabelle

    2013-01-01

    Limb-girdle muscular dystrophy 2D (LGMD2D) is an inherited myogenic disorder belonging to the group of muscular dystrophies. Sgca-null mouse is a knock-out model of LGMD2D. Little is known about cardiac phenotype characterization in this model at different ages. We conducted a prospective study to characterize cardiac sgca-null mice phenotype using high resolution Doppler echocardiography at different ages. Conventional echocardiography was performed on anesthetised mice using a Vevo 770 (Visualsonics) with 30 MHz cardiac probe. Wild Type (WT) and sgca-null mice were scanned at 13, 15 and 17 months. From M-mode, we measured interventricular septal (IVS) wall thickness, posterior wall (PW) thickness, and end-left ventricular diameter in systolic and diastolic. From the above parameters, we calculated left ventricular (LV) shortening fraction (SF), LV ejection fraction (EF) and LV mass. At age 13 months, PW diastolic thickness was increased in sgca-null mice (0.89±0.14 mm vs 0.73±0.2 mm; P=0.020) and LV mass was higher in sgca-null mice (LV mass 205.2 mg vs 143 mg; P=0.001). We found also dilation of the LV (LVEDD: 4.84 mm vs 4.29 mm; P=0.019) in sgca-null mice. At age 15 months, dilation of the LV (LVEDD: 4.86 mm vs 4 mm; P=0.05) with an increase of the LV mass (165.7 mg vs 127.12; P=0.03) are found in sgca-null mice. At age 17 months, we found a decrease of the PW thickening (17% vs 30%; P=0.036). This work provides echocardiographic insights for the assessment of pharmaceutical therapies in sgca-null mice. PMID:24416486

  18. Hepatic effects of a methionine-choline-deficient diet in hepatocyte RXR{alpha}-null mice

    SciTech Connect

    Gyamfi, Maxwell Afari; Tanaka, Yuji; He Lin; Klaassen, Curtis D.; Wan, Y.-J.Y.

    2009-01-15

    Retinoid X receptor-{alpha} (RXR{alpha}) is an obligate partner for several nuclear hormone receptors that regulate important physiological processes in the liver. In this study the impact of hepatocyte RXR{alpha} deficiency on methionine and choline deficient (MCD) diet-induced steatosis, oxidative stress, inflammation, and hepatic transporters gene expression were examined. The mRNA of sterol regulatory element-binding protein (SREBP)-regulated genes, important for lipid synthesis, were not altered in wild type (WT) mice, but were increased 2.0- to 5.4-fold in hepatocyte RXR{alpha}-null (H-RXR{alpha}-null) mice fed a MCD diet for 14 days. Furthermore, hepatic mRNAs and proteins essential for fatty acid {beta}-oxidation were not altered in WT mice, but were decreased in the MCD diet-fed H-RXR{alpha}-null mice, resulting in increased hepatic free fatty acid levels. Cyp2e1 enzyme activity and lipid peroxide levels were induced only in MCD-fed WT mice. In contrast, hepatic mRNA levels of pro-inflammatory factors were increased only in H-RXR{alpha}-null mice fed the MCD diet. Hepatic uptake transporters Oatp1a1 and Oatp1b2 mRNA levels were decreased in WT mice fed the MCD diet, whereas the efflux transporter Mrp4 was increased. However, in the H-RXR{alpha}-null mice, the MCD diet only moderately decreased Oatp1a1 and induced both Oatp1a4 and Mrp4 gene expression. Whereas the MCD diet increased serum bile acid levels and alkaline phosphatase activity in both WT and H-RXR{alpha}-null mice, serum ALT levels were induced (2.9-fold) only in the H-RXR{alpha}-null mice. In conclusion, these data suggest a critical role for RXR{alpha} in hepatic fatty acid homeostasis and protection against MCD-induced hepatocyte injury.

  19. Organic anion transporting polypeptide 1a1 null mice are sensitive to cholestatic liver injury.

    PubMed

    Zhang, Youcai; Csanaky, Iván L; Cheng, Xingguo; Lehman-McKeeman, Lois D; Klaassen, Curtis D

    2012-06-01

    Organic anion transporting polypeptide 1a1 (Oatp1a1) is predominantly expressed in livers of mice and is thought to transport bile acids (BAs) from blood into liver. Because Oatp1a1 expression is markedly decreased in mice after bile duct ligation (BDL). We hypothesized that Oatp1a1-null mice would be protected against liver injury during BDL-induced cholestasis due largely to reduced hepatic uptake of BAs. To evaluate this hypothesis, BDL surgeries were performed in both male wild-type (WT) and Oatp1a1-null mice. At 24 h after BDL, Oatp1a1-null mice showed higher serum alanine aminotransferase levels and more severe liver injury than WT mice, and all Oatp1a1-null mice died within 4 days after BDL, whereas all WT mice survived. At 24 h after BDL, surprisingly Oatp1a1-null mice had higher total BA concentrations in livers than WT mice, suggesting that loss of Oatp1a1 did not prevent BA accumulation in the liver. In addition, secondary BAs dramatically increased in serum of Oatp1a1-null BDL mice but not in WT BDL mice. Oatp1a1-null BDL mice had similar basolateral BA uptake (Na(+)-taurocholate cotransporting polypeptide and Oatp1b2) and BA-efflux (multidrug resistance-associated protein [Mrp]-3, Mrp4, and organic solute transporter α/β) transporters, as well as BA-synthetic enzyme (Cyp7a1) in livers as WT BDL mice. Hepatic expression of small heterodimer partner Cyp3a11, Cyp4a14, and Nqo1, which are target genes of farnesoid X receptor, pregnane X receptor, peroxisome proliferator-activated receptor alpha, and NF-E2-related factor 2, respectively, were increased in WT BDL mice but not in Oatp1a1-null BDL mice. These results demonstrate that loss of Oatp1a1 function exacerbates cholestatic liver injury in mice and suggest that Oatp1a1 plays a unique role in liver adaptive responses to obstructive cholestasis.

  20. Organic Anion Transporting Polypeptide 1a1 Null Mice Are Sensitive to Cholestatic Liver Injury

    PubMed Central

    Zhang, Youcai; Csanaky, Iván L.; Cheng, Xingguo; Lehman-McKeeman, Lois D.; Klaassen, Curtis D.

    2012-01-01

    Organic anion transporting polypeptide 1a1 (Oatp1a1) is predominantly expressed in livers of mice and is thought to transport bile acids (BAs) from blood into liver. Because Oatp1a1 expression is markedly decreased in mice after bile duct ligation (BDL). We hypothesized that Oatp1a1-null mice would be protected against liver injury during BDL-induced cholestasis due largely to reduced hepatic uptake of BAs. To evaluate this hypothesis, BDL surgeries were performed in both male wild-type (WT) and Oatp1a1-null mice. At 24 h after BDL, Oatp1a1-null mice showed higher serum alanine aminotransferase levels and more severe liver injury than WT mice, and all Oatp1a1-null mice died within 4 days after BDL, whereas all WT mice survived. At 24 h after BDL, surprisingly Oatp1a1-null mice had higher total BA concentrations in livers than WT mice, suggesting that loss of Oatp1a1 did not prevent BA accumulation in the liver. In addition, secondary BAs dramatically increased in serum of Oatp1a1-null BDL mice but not in WT BDL mice. Oatp1a1-null BDL mice had similar basolateral BA uptake (Na+-taurocholate cotransporting polypeptide and Oatp1b2) and BA-efflux (multidrug resistance–associated protein [Mrp]-3, Mrp4, and organic solute transporter α/β) transporters, as well as BA-synthetic enzyme (Cyp7a1) in livers as WT BDL mice. Hepatic expression of small heterodimer partner Cyp3a11, Cyp4a14, and Nqo1, which are target genes of farnesoid X receptor, pregnane X receptor, peroxisome proliferator-activated receptor alpha, and NF-E2-related factor 2, respectively, were increased in WT BDL mice but not in Oatp1a1-null BDL mice. These results demonstrate that loss of Oatp1a1 function exacerbates cholestatic liver injury in mice and suggest that Oatp1a1 plays a unique role in liver adaptive responses to obstructive cholestasis. PMID:22461449

  1. Abnormal Mammary Development in 129:STAT1-Null Mice is Stroma-Dependent.

    PubMed

    Chen, Jane Q; Mori, Hidetoshi; Cardiff, Robert D; Trott, Josephine F; Hovey, Russell C; Hubbard, Neil E; Engelberg, Jesse A; Tepper, Clifford G; Willis, Brandon J; Khan, Imran H; Ravindran, Resmi K; Chan, Szeman R; Schreiber, Robert D; Borowsky, Alexander D

    2015-01-01

    Female 129:Stat1-null mice (129S6/SvEvTac-Stat1(tm1Rds) homozygous) uniquely develop estrogen-receptor (ER)-positive mammary tumors. Herein we report that the mammary glands (MG) of these mice have altered growth and development with abnormal terminal end buds alongside defective branching morphogenesis and ductal elongation. We also find that the 129:Stat1-null mammary fat pad (MFP) fails to sustain the growth of 129S6/SvEv wild-type and Stat1-null epithelium. These abnormalities are partially reversed by elevated serum progesterone and prolactin whereas transplantation of wild-type bone marrow into 129:Stat1-null mice does not reverse the MG developmental defects. Medium conditioned by 129:Stat1-null epithelium-cleared MFP does not stimulate epithelial proliferation, whereas it is stimulated by medium conditioned by epithelium-cleared MFP from either wild-type or 129:Stat1-null females having elevated progesterone and prolactin. Microarrays and multiplexed cytokine assays reveal that the MG of 129:Stat1-null mice has lower levels of growth factors that have been implicated in normal MG growth and development. Transplanted 129:Stat1-null tumors and their isolated cells also grow slower in 129:Stat1-null MG compared to wild-type recipient MG. These studies demonstrate that growth of normal and neoplastic 129:Stat1-null epithelium is dependent on the hormonal milieu and on factors from the mammary stroma such as cytokines. While the individual or combined effects of these factors remains to be resolved, our data supports the role of STAT1 in maintaining a tumor-suppressive MG microenvironment.

  2. Abnormal Mammary Development in 129:STAT1-Null Mice is Stroma-Dependent

    PubMed Central

    Cardiff, Robert D.; Trott, Josephine F.; Hovey, Russell C.; Hubbard, Neil E.; Engelberg, Jesse A.; Tepper, Clifford G.; Willis, Brandon J.; Khan, Imran H.; Ravindran, Resmi K.; Chan, Szeman R.; Schreiber, Robert D.; Borowsky, Alexander D.

    2015-01-01

    Female 129:Stat1-null mice (129S6/SvEvTac-Stat1tm1Rds homozygous) uniquely develop estrogen-receptor (ER)-positive mammary tumors. Herein we report that the mammary glands (MG) of these mice have altered growth and development with abnormal terminal end buds alongside defective branching morphogenesis and ductal elongation. We also find that the 129:Stat1-null mammary fat pad (MFP) fails to sustain the growth of 129S6/SvEv wild-type and Stat1-null epithelium. These abnormalities are partially reversed by elevated serum progesterone and prolactin whereas transplantation of wild-type bone marrow into 129:Stat1-null mice does not reverse the MG developmental defects. Medium conditioned by 129:Stat1-null epithelium-cleared MFP does not stimulate epithelial proliferation, whereas it is stimulated by medium conditioned by epithelium-cleared MFP from either wild-type or 129:Stat1-null females having elevated progesterone and prolactin. Microarrays and multiplexed cytokine assays reveal that the MG of 129:Stat1-null mice has lower levels of growth factors that have been implicated in normal MG growth and development. Transplanted 129:Stat1-null tumors and their isolated cells also grow slower in 129:Stat1-null MG compared to wild-type recipient MG. These studies demonstrate that growth of normal and neoplastic 129:Stat1-null epithelium is dependent on the hormonal milieu and on factors from the mammary stroma such as cytokines. While the individual or combined effects of these factors remains to be resolved, our data supports the role of STAT1 in maintaining a tumor-suppressive MG microenvironment. PMID:26075897

  3. Gender- and region-specific alterations in bone metabolism in Scarb1-null female mice.

    PubMed

    Martineau, Corine; Martin-Falstrault, Louise; Brissette, Louise; Moreau, Robert

    2014-08-01

    A positive correlation between plasma levels of HDL and bone mass has been reported by epidemiological studies. As scavenger receptor class B, type I (SR-BI), the gene product of Scarb1, is known to regulate HDL metabolism, we recently characterized bone metabolism in Scarb1-null mice. These mice display high femoral bone mass associated with enhanced bone formation. As gender differences have been reported in HDL metabolism and SR-BI function, we investigated gender-specific bone alterations in Scarb1-null mice by microtomography and histology. We found 16% greater relative bone volume and 39% higher bone formation rate in the vertebrae from 2-month-old Scarb1-null females. No such alteration was seen in males, indicating gender- and region-specific differences in skeletal phenotype. Total and HDL-associated cholesterol levels, as well as ACTH plasma levels, were increased in both Scarb1-null genders, the latter being concurrent to impaired corticosterone response to fasting. Plasma levels of estradiol did not differ between null and WT females, suggesting that the estrogen metabolism alteration is not relevant to the higher vertebral bone mass in female Scarb1-null mice. Constitutively, high plasma levels of leptin along with 2.5-fold increase in its expression in white adipose tissue were measured in female Scarb1-null mice only. In vitro exposure of bone marrow stromal cells to ACTH and leptin promoted osteoblast differentiation as evidenced by increased gene expression of osterix and collagen type I alpha. Our results suggest that hyperleptinemia may account for the gender-specific high bone mass seen in the vertebrae of female Scarb1-null mice.

  4. Antagonism of angiotensin 1–7 prevents the therapeutic effects of recombinant human ACE2

    PubMed Central

    Patel, Vaibhav B.; Takawale, Abhijit; Ramprasath, Tharmarajan; Das, Subhash K.; Basu, Ratnadeep; Grant, Maria B.; Hall, David A.; Kassiri, Zamaneh

    2015-01-01

    Activation of the angiotensin 1–7/Mas receptor (MasR) axis counteracts angiotensin II (Ang II)-mediated cardiovascular disease. Recombinant human angiotensin-converting enzyme 2 (rhACE2) generates Ang 1–7 from Ang II. We hypothesized that the therapeutic effects of rhACE2 are dependent on Ang 1–7 action. Wild type male C57BL/6 mice (10–12 weeks old) were infused with Ang II (1.5 mg/kg/d) and treated with rhACE2 (2 mg/kg/d). The Ang 1–7 antagonist, A779 (200 ng/kg/min), was administered to a parallel group of mice. rhACE2 prevented Ang II-induced hypertrophy and diastolic dysfunction while A779 prevented these beneficial effects and precipitated systolic dysfunction. rhACE2 effectively antagonized Ang II-mediated myocardial fibrosis which was dependent on the action of Ang 1–7. Myocardial oxidative stress and matrix metalloproteinase 2 activity was further increased by Ang 1–7 inhibition even in the presence of rhACE2. Activation of Akt and endothelial nitric oxide synthase (eNOS) by rhACE2 were suppressed by the antagonism of Ang 1–7 while the activation of pathological signaling pathways was maintained. Blocking Ang 1–7 action prevents the therapeutic effects of rhACE2 in the setting of elevated Ang II culminating in systolic dysfunction. These results highlight a key cardioprotective role of Ang 1–7, and increased Ang 1–7 action represents a potential therapeutic strategy for cardiovascular diseases. PMID:25874965

  5. Abnormal Activation of BMP Signaling Causes Myopathy in Fbn2 Null Mice.

    PubMed

    Sengle, Gerhard; Carlberg, Valerie; Tufa, Sara F; Charbonneau, Noe L; Smaldone, Silvia; Carlson, Eric J; Ramirez, Francesco; Keene, Douglas R; Sakai, Lynn Y

    2015-06-01

    Fibrillins are large extracellular macromolecules that polymerize to form the backbone structure of connective tissue microfibrils. Mutations in the gene for fibrillin-1 cause the Marfan syndrome, while mutations in the gene for fibrillin-2 cause Congenital Contractural Arachnodactyly. Both are autosomal dominant disorders, and both disorders affect musculoskeletal tissues. Here we show that Fbn2 null mice (on a 129/Sv background) are born with reduced muscle mass, abnormal muscle histology, and signs of activated BMP signaling in skeletal muscle. A delay in Myosin Heavy Chain 8, a perinatal myosin, was found in Fbn2 null forelimb muscle tissue, consistent with the notion that muscle defects underlie forelimb contractures in these mice. In addition, white fat accumulated in the forelimbs during the early postnatal period. Adult Fbn2 null mice are already known to demonstrate persistent muscle weakness. Here we measured elevated creatine kinase levels in adult Fbn2 null mice, indicating ongoing cycles of muscle injury. On a C57Bl/6 background, Fbn2 null mice showed severe defects in musculature, leading to neonatal death from respiratory failure. These new findings demonstrate that loss of fibrillin-2 results in phenotypes similar to those found in congenital muscular dystrophies and that FBN2 should be considered as a candidate gene for recessive congenital muscular dystrophy. Both in vivo and in vitro evidence associated muscle abnormalities and accumulation of white fat in Fbn2 null mice with abnormally activated BMP signaling. Genetic rescue of reduced muscle mass and accumulation of white fat in Fbn2 null mice was accomplished by deleting a single allele of Bmp7. In contrast to other reports that activated BMP signaling leads to muscle hypertrophy, our findings demonstrate the exquisite sensitivity of BMP signaling to the fibrillin-2 extracellular environment during early postnatal muscle development. New evidence presented here suggests that fibrillin-2 can

  6. Secreted Protein Acidic and Rich in Cysteine (SPARC)-Null Mice Exhibit More Uniform Outflow

    PubMed Central

    Swaminathan, Swarup S.; Oh, Dong-Jin; Kang, Min Hyung; Ren, Ruiyi; Jin, Rui; Gong, Haiyan; Rhee, Douglas J.

    2013-01-01

    Purpose. Secreted protein acidic and rich in cysteine (SPARC) is a matricellular protein known to regulate extracellular matrix (ECM) in many tissues and is highly expressed in trabecular meshwork (TM). SPARC-null mice have a 15% to 20% decrease in intraocular pressure (IOP) compared to wild-type (WT) mice. We hypothesized that mouse aqueous outflow is segmental, and that transgenic deletion of SPARC causes a more uniform pattern that correlates with IOP and TM morphology. Methods. Eyes of C57BL6-SV129 WT and SPARC-null mice were injected with fluorescent microbeads, which were also passively exposed to freshly enucleated eyes. Confocal and electron microscopy were performed. Percentage effective filtration length (PEFL) was calculated as PEFL = FL/TL × 100%, where TL = total length and FL = filtration length. IOP was measured by rebound tonometry. Results. Passive microbead affinity for WT and SPARC-null ECM did not differ. Segmental flow was observed in the mouse eye. SPARC-null mice had a 23% decrease in IOP. PEFL increased in SPARC-null (70.61 ± 11.36%) versus WT mice (54.68 ± 9.95%, P < 0.005; n = 11 pairs), and PEFL and IOP were negatively correlated (R2 = 0.72, n = 10 pairs). Morphologically, TM of high-tracer regions had increased separation between beams compared to low-tracer regions. Collagen fibril diameter decreased in SPARC-null (28.272 nm) versus WT tissue (34.961 nm, P < 0.0005; n = 3 pairs). Conclusions. Aqueous outflow in mice is segmental. SPARC-null mice demonstrated a more uniform outflow pattern and decreased collagen fibril diameter. Areas of high flow had less compact juxtacanalicular connective tissue ECM, and IOP was inversely correlated with PEFL. Our data show a correlation between morphology, aqueous outflow, and IOP, indicating a modulatory role of SPARC in IOP regulation. PMID:23422826

  7. Increased Oxidative Stress Impairs Adipose Tissue Function in Sphingomyelin Synthase 1 Null Mice

    PubMed Central

    Nishimura, Naotaka; Gotoh, Tomomi; Watanabe, Ken; Ikeda, Kazutaka; Garan, Yohei; Taguchi, Ryo; Node, Koichi; Okazaki, Toshiro; Oike, Yuichi

    2013-01-01

    Sphingomyelin synthase 1 (SMS1) catalyzes the conversion of ceramide to sphingomyelin. Here, we found that SMS1 null mice showed lipodystrophic phenotype. Mutant mice showed up-regulation of plasma triglyceride concentrations accompanied by reduction of white adipose tissue (WAT) as they aged. Lipoprotein lipase (LPL) activity was severely reduced in mutant mice. In vivo analysis indicated that fatty acid uptake in WAT but not in liver decreased in SMS1 null compared to wild-type mice. In vitro analysis using cultured cell revealed that SMS1 depletion reduced fatty acid uptake. Proteins extracted from WAT of mutant mice were severely modified by oxidative stress, and up-regulation of mRNAs related to apoptosis, redox adjustment, mitochondrial stress response and mitochondrial biogenesis was observed. ATP content of WAT was reduced in SMS1 null mice. Blue native gel analysis indicated that accumulation of mitochondrial respiratory chain complexes was reduced. These results suggest that WAT of SMS1 null mice is severely damaged by oxidative stress and barely functional. Indeed, mutant mice treated with the anti-oxidant N-acetyl cysteine (NAC) showed partial recovery of lipodystrophic phenotypes together with normalized plasma triglyceride concentrations. Altogether, our data suggest that SMS1 is crucial to control oxidative stress in order to maintain WAT function. PMID:23593476

  8. Urinary metabolomics in Fxr-null mice reveals activated adaptive metabolic pathways upon bile acid challenge.

    PubMed

    Cho, Joo-Youn; Matsubara, Tsutomu; Kang, Dong Wook; Ahn, Sung-Hoon; Krausz, Kristopher W; Idle, Jeffrey R; Luecke, Hans; Gonzalez, Frank J

    2010-05-01

    Farnesoid X receptor (FXR) is a nuclear receptor that regulates genes involved in synthesis, metabolism, and transport of bile acids and thus plays a major role in maintaining bile acid homeostasis. In this study, metabolomic responses were investigated in urine of wild-type and Fxr-null mice fed cholic acid, an FXR ligand, using ultra-performance liquid chromatography (UPLC) coupled with electrospray time-of-flight mass spectrometry (TOFMS). Multivariate data analysis between wild-type and Fxr-null mice on a cholic acid diet revealed that the most increased ions were metabolites of p-cresol (4-methylphenol), corticosterone, and cholic acid in Fxr-null mice. The structural identities of the above metabolites were confirmed by chemical synthesis and by comparing retention time (RT) and/or tandem mass fragmentation patterns of the urinary metabolites with the authentic standards. Tauro-3alpha,6,7alpha,12alpha-tetrol (3alpha,6,7alpha,12alpha-tetrahydroxy-5beta-cholestan-26-oyltaurine), one of the most increased metabolites in Fxr-null mice on a CA diet, is a marker for efficient hydroxylation of toxic bile acids possibly through induction of Cyp3a11. A cholestatic model induced by lithocholic acid revealed that enhanced expression of Cyp3a11 is the major defense mechanism to detoxify cholestatic bile acids in Fxr-null mice. These results will be useful for identification of biomarkers for cholestasis and for determination of adaptive molecular mechanisms in cholestasis.

  9. Dmp1 Null Mice Develop a Unique Osteoarthritis-like Phenotype

    PubMed Central

    Zhang, Qi; Lin, Shuxian; Liu, Ying; Yuan, Baozhi; Harris, Steph E; Feng, Jian Q.

    2016-01-01

    Patients with hypophosphatemia rickets (including DMP1 mutations) develop severe osteoarthritis (OA), although the mechanism is largely unknown. In this study, we first identified the expression of DMP1 in hypertrophic chondrocytes using immunohistochemistry (IHC) and X-gal analysis of Dmp1-knockout-lacZ-knockin heterozygous mice. Next, we characterized the OA-like phenotype in Dmp1 null mice from 7-week-old to one-year-old using multiple techniques, including X-ray, micro-CT, H&E staining, Goldner staining, scanning electronic microscopy, IHC assays, etc. We found a classical OA-like phenotype in Dmp1 null mice such as articular cartilage degradation, osteophyte formation, and subchondral osteosclerosis. These Dmp1 null mice also developed unique pathological changes, including a biphasic change in their articular cartilage from the initial expansion of hypertrophic chondrocytes at the age of 1-month to a quick diminished articular cartilage layer at the age of 3-months. Further, these null mice displayed severe enlarged knees and poorly formed bone with an expanded osteoid area. To address whether DMP1 plays a direct role in the articular cartilage, we deleted Dmp1 specifically in hypertrophic chondrocytes by crossing the Dmp1-loxP mice with Col X Cre mice. Interestingly, these conditional knockout mice didn't display notable defects in either the articular cartilage or the growth plate. Because of the hypophosphatemia remained in the entire life span of the Dmp1 null mice, we also investigated whether a high phosphate diet would improve the OA-like phenotype. A 8-week treatment of a high phosphate diet significantly rescued the OA-like defect in Dmp1 null mice, supporting the critical role of phosphate homeostasis in maintaining the healthy joint morphology and function. Taken together, this study demonstrates a unique OA-like phenotype in Dmp1 null mice, but a lack of the direct impact of DMP1 on chondrogenesis. Instead, the regulation of phosphate homeostasis

  10. Dmp1 Null Mice Develop a Unique Osteoarthritis-like Phenotype.

    PubMed

    Zhang, Qi; Lin, Shuxian; Liu, Ying; Yuan, Baozhi; Harris, Steph E; Feng, Jian Q

    2016-01-01

    Patients with hypophosphatemia rickets (including DMP1 mutations) develop severe osteoarthritis (OA), although the mechanism is largely unknown. In this study, we first identified the expression of DMP1 in hypertrophic chondrocytes using immunohistochemistry (IHC) and X-gal analysis of Dmp1-knockout-lacZ-knockin heterozygous mice. Next, we characterized the OA-like phenotype in Dmp1 null mice from 7-week-old to one-year-old using multiple techniques, including X-ray, micro-CT, H&E staining, Goldner staining, scanning electronic microscopy, IHC assays, etc. We found a classical OA-like phenotype in Dmp1 null mice such as articular cartilage degradation, osteophyte formation, and subchondral osteosclerosis. These Dmp1 null mice also developed unique pathological changes, including a biphasic change in their articular cartilage from the initial expansion of hypertrophic chondrocytes at the age of 1-month to a quick diminished articular cartilage layer at the age of 3-months. Further, these null mice displayed severe enlarged knees and poorly formed bone with an expanded osteoid area. To address whether DMP1 plays a direct role in the articular cartilage, we deleted Dmp1 specifically in hypertrophic chondrocytes by crossing the Dmp1-loxP mice with Col X Cre mice. Interestingly, these conditional knockout mice didn't display notable defects in either the articular cartilage or the growth plate. Because of the hypophosphatemia remained in the entire life span of the Dmp1 null mice, we also investigated whether a high phosphate diet would improve the OA-like phenotype. A 8-week treatment of a high phosphate diet significantly rescued the OA-like defect in Dmp1 null mice, supporting the critical role of phosphate homeostasis in maintaining the healthy joint morphology and function. Taken together, this study demonstrates a unique OA-like phenotype in Dmp1 null mice, but a lack of the direct impact of DMP1 on chondrogenesis. Instead, the regulation of phosphate homeostasis

  11. Nitrosative stress elicited by nNOSµ delocalization inhibits muscle force in dystrophin-null mice.

    PubMed

    Li, Dejia; Yue, Yongping; Lai, Yi; Hakim, Chady H; Duan, Dongsheng

    2011-01-01

    The mechanism of force reduction is not completely understood in Duchenne muscular dystrophy (DMD), a dystrophin-deficient lethal disease. Nitric oxide regulates muscle force. Interestingly, neuronal nitric oxide synthase µ (nNOSµ), a major source of muscle nitric oxide, is lost from the sarcolemma in DMD muscle. We hypothesize that nNOSµ delocalization contributes to force reduction in DMD. To test this hypothesis, we generated dystrophin/nNOSµ double knockout mice. Genetic elimination of nNOSµ significantly enhanced force in dystrophin-null mice. Pharmacological inhibition of nNOS yielded similar results. To further test our hypothesis, we studied δ-sarcoglycan-null mice, a model of limb-girdle muscular dystrophy. These mice had minimal sarcolemmal nNOSµ delocalization and muscle force was less compromised. Annihilation of nNOSµ did not improve their force either. To determine whether nNOSµ delocalization itself inhibited force, we corrected muscle disease in dystrophin-null mice with micro-dystrophins that either restored or did not restore sarcolemmal nNOSµ. Similar muscle force was obtained irrespective of nNOSµ localization. Additional studies suggest that nNOSµ delocalization selectively inhibits muscle force in dystrophin-null mice via nitrosative stress. In summary, we have demonstrated for the first time that nitrosative stress elicited by nNOSµ delocalization is an important mechanism underlying force loss in DMD.

  12. Stimulation of Sigma-1 Receptor Ameliorates Depressive-like Behaviors in CaMKIV Null Mice.

    PubMed

    Moriguchi, Shigeki; Sakagami, Hiroyuki; Yabuki, Yasushi; Sasaki, Yuzuru; Izumi, Hisanao; Zhang, Chen; Han, Feng; Fukunaga, Kohji

    2015-12-01

    Sigma-1 receptor (Sig-1R) is a molecular chaperone regulating calcium efflux from the neuronal endoplasmic reticulum to the mitochondria. Calcium/calmodulin-dependent protein kinase IV (CaMKIV) null mice exhibit depressive-like behaviors and impaired neurogenesis as assessed by bromodeoxyuridine (BrdU) incorporation into newborn cells of the hippocampal dentate gyrus (DG). Here, we demonstrate that chronic stimulation of Sig-1R by treatment with the agonist SA4503 or the SSRI fluvoxamine for 14 days improves depressive-like behaviors in CaMKIV null mice. By contrast, treatment with paroxetine, which lacks affinity for Sig-1R, did not alter these behaviors. Reduced numbers of BrdU-positive cells and decreased brain-derived neurotrophic factor (BDNF) mRNA expression and protein kinase B (Akt; Ser-473) phosphorylation seen in the DG of CaMKIV null mice were significantly rescued by chronic Sig-1R stimulation. Interestingly, reduced ATP production observed in the DG of CaMKIV null mice was improved by chronic Sig-1R stimulation. Such stimulation also improved hippocampal long-term potentiation (LTP) induction and maintenance, which are impaired in the DG of CaMKIV null mice. LTP rescue was closely associated with both increases in calcium/calmodulin-dependent protein kinase II (CaMKII) autophosphorylation and GluA1 (Ser-831) phosphorylation. Taken together, Sig-1R stimulation by SA4503 or fluvoxamine treatment increased hippocampal neurogenesis, which is closely associated with amelioration of depressive-like behaviors in CaMKIV null mice.

  13. Loss of FAS/FASL signaling does not reduce apoptosis in Sharpin null mice.

    PubMed

    Potter, Christopher S; Silva, Kathleen A; Kennedy, Victoria E; Stearns, Timothy M; HogenEsch, Harm; Sundberg, John P

    2017-01-17

    Mice with mutations in SHANK-associated RH domain interactor (Sharpin) develop a hypereosinophilic auto-inflammatory disease known as chronic proliferative dermatitis. Affected mice have increased apoptosis in the keratinocytes of the skin, esophagus, and forestomach driven by extrinsic TNF receptor mediated apoptotic signaling pathways. FAS receptor signaling is an extrinsic apoptotic signaling mechanism frequently involved in inflammatory skin diseases. Compound mutations in Sharpin and Fas or Fasl were created to determine if these death domain proteins influenced the cutaneous phenotype in Sharpin null mice. Both Sharpin/Fas and Sharpin/Fasl compound mutant mice developed an auto-inflammatory phenotype similar to that seen in Sharpin null mice indicating that initiation of apoptosis by FAS signaling is likely not involved in the pathogenesis of this disease. This article is protected by copyright. All rights reserved.

  14. Incisional wound healing in transforming growth factor-beta1 null mice.

    PubMed

    Koch, R M; Roche, N S; Parks, W T; Ashcroft, G S; Letterio, J J; Roberts, A B

    2000-01-01

    Expression of endogenous transforming growth factor-beta1 is reduced in many animal models of impaired wound healing, and addition of exogenous transforming growth factor-beta has been shown to improve healing. To test the hypothesis that endogenous transforming growth factor-beta1 is essential for normal wound repair, we have studied wound healing in mice in which the transforming growth factor-beta1 gene has been deleted by homologous recombination. No perceptible differences were observed in wounds made in 3-10-day-old neonatal transforming growth factor-beta1 null mice compared to wild-type littermates. To preclude interference from maternally transferred transforming growth factor-beta1, cutaneous wounds were also made on the backs of 30-day-old transforming growth factor-beta1 null and littermate control mice treated with rapamycin, which extends their lifetime and suppresses the inflammatory response characteristic of the transforming growth factor-beta1 null mice. Again, no impairment in healing was seen in transforming growth factor-beta1 null mice. Instead these wounds showed an overall reduction in the amount of granulation tissue and an increased rate of epithelialization compared to littermate controls. Our data suggest that release of transforming growth factor-beta1 from degranulating platelets or secretion by infiltrating macrophages and fibroblasts is not critical to initiation or progression of tissue repair and that endogenous transforming growth factor-beta1 may actually function to increase inflammation and retard wound closure.

  15. Evidence of altered inhibition in layer V pyramidal neurons from neocortex of Kcna1-null mice.

    PubMed

    van Brederode, J F; Rho, J M; Cerne, R; Tempel, B L; Spain, W J

    2001-01-01

    Mice lacking the potassium channel subunit KCNA1 exhibit a severe epileptic phenotype beginning at an early postnatal age. The precise cellular physiological substrates for these seizures are unclear, as is the site of origin. Since KCNA1 mRNA in normal mice is expressed in the neocortex, we asked whether neurons in the neocortex of three to four week-old Kcna1-null mutants exhibit evidence of hyperexcitability. Layer V pyramidal neurons were directly visualized in brain slices with infrared differential-interference contrast microscopy and evaluated with cellular electrophysiological techniques. There were no significant differences in intrinsic membrane properties and action potential shape between Kcna1-null and wild-type mice, consistent with previous findings in hippocampal slice recordings. However, the frequency of spontaneous post-synaptic currents was significantly higher in Kcna1-null compared to wild-type mice. The frequency of spontaneous inhibitory post-synaptic currents and miniature (action-potential-independent) inhibitory post-synaptic currents was also significantly higher in Kcna1-null compared to wild-type mice. However, the frequency of spontaneous and miniature excitatory post-synaptic currents was not different in these two groups of animals. Comparison of the amplitude and kinetics of miniature inhibitory and excitatory post-synaptic currents revealed differences in amplitude, rise time and half-width between Kcna1-null and wild-type mice. Our data indicate that the inhibitory drive onto layer V pyramidal neurons is increased in Kcna1 knockout mice, either directly through an increased spontaneous release of GABA from presynaptic terminals contacting layer V pyramidal neurons, or an enhanced excitatory synaptic input to inhibitory interneurons.

  16. Wound Healing Characteristics of ICAM-1 Null Mice Devoid of All Isoforms of ICAM-1

    PubMed Central

    Gay, Andre N.; Mushin, Oren P.; Lazar, David A.; Naik-Mathuria, Bindi J.; Yu, Ling; Gobin, Andre; Smith, C. Wayne; Olutoye, Oluyinka O.

    2011-01-01

    Background Intercellular Adhesion Molecule-1 (ICAM-1) permits leukocyte-endothelial adhesion and transmigration during inflammation. Membrane-bound ICAM-1 knockout mice have been used to understand this molecule’s role in wound-healing, but expressed spliced isoforms of ICAM-1 may have impacted results. We aimed to characterize wound-healing in an ICAM-1 null model devoid of all ICAM-1 isoforms. Methods Full-thickness 8-mm wounds were created on C57/BL6 wild-type (n=24) and ICAM-1 null (n=24) mice. Wound area was calculated using daily photographs. Histologic samples were harvested on post-operative Days 1,3,7, and 14. Wound margins were evaluated for mRNA expression of 13 inflammatory cytokines. A separate group of wild-type and ICAM-1 null mice (n=24) received full-thickness incisions with tensiometry measured at Day 14. Separately, complete blood counts were measured in unwounded wild-type (n=4) and ICAM-1 null mice (n=4). Results Wound-closure was significantly delayed in ICAM-1 null mice through Day 7 by gross and histologic measurement. mRNA expression of VEGF-A was increased in ICAM-1 null mice on Day 3, although no increase in VEGF-A was observed in the wound bed by immunohistochemistry. ICAM-1 null wounds demonstrated higher stiffness upon Day 14 tensiometry compared to the wild-type (1880 ± 926 kPa vs. 478 ± 117 kPa;p<0.01), and had higher counts of white blood cells (10,009 vs. 5,720 cells/microliter,p<0.05), neutrophils (2,130 vs. 630 cells/microliter,p<0.01), and lymphocytes (7,130 vs. 4,740 cells/microliter, p<0.05). Conclusions ICAM-1 null mice demonstrate delayed wound-healing and decreased wound elasticity compared to wild-type controls. This lag, however, was less than observed in earlier membrane-bound ICAM-1 knockouts, suggesting that other ICAM-1 isoforms may promote delayed wound-healing. PMID:21872884

  17. Enhanced Autophagy in Polycystic Kidneys of AQP11 Null Mice

    PubMed Central

    Tanaka, Yasuko; Watari, Mayumi; Saito, Tatsuya; Morishita, Yoshiyuki; Ishibashi, Kenichi

    2016-01-01

    Aquaporin-11 (AQP11) is an intracellular water channel expressed at the endoplasmic reticulum (ER) of the proximal tubule. Its gene disruption in mice leads to intracellular vacuole formation at one week and the subsequent development of polycystic kidneys by three weeks. As the damaged proximal tubular cells with intracellular vacuoles form cysts later, we postulated that autophagy may play a role in the cyst formation and examined autophagy activity before and after cyst development in AQP11(−/−) kidneys. PCR analysis showed the increased expression of the transcript encoding LC3 (Map1lc3b) as well as other autophagy-related genes in AQP11(−/−) mice. Using green fluorescent protein (GFP)-LC3 transgenic mice and AQP11(−/−) mice, we found that the number of GFP-LC3–positive puncta was increased in the proximal tubule of AQP11(−/−) mice before the cyst formation. Interestingly, they were also observed in the cyst-lining epithelial cell. Further PCR analyses revealed the enhanced expression of apoptosis-related and ER stress–related caspase genes before and after the cyst formation, which may cause the enhanced autophagy. These results suggest the involvement of autophagy in the development and maintenance of kidney cysts in AQP11(−/−) mice. PMID:27916883

  18. Growth defect in Grg5 null mice is associated with reduced Ihh signaling in growth plates.

    PubMed

    Wang, Wen-Fang; Wang, You-Gan; Reginato, Anthony M; Plotkina, Sofiya; Gridley, Thomas; Olsen, Bjorn R

    2002-05-01

    Gene-targeted disruption of Grg5, a mouse homologue of Drosophila groucho (gro), results in postnatal growth retardation in mice. The growth defect, most striking in approximately half of the Grg5 null mice, occurs during the first 4-5 weeks of age, but most mice recover retarded growth later. We used the nonlinear mixed-effects model to fit the growth data of wild-type, heterozygous, and Grg5 null mice. On the basis of preliminary evidence suggesting an interaction between Grg5 and the transcription factor Cbfa1/Runx2, critical for skeletal development, we further investigated the skeleton in the mice. A long bone growth plate defect was identified, which included shorter zones of proliferative and hypertrophic chondrocytes and decreased trabecular bone formation. This decreased trabecular bone formation is likely caused by a reduced recruitment of osteoblasts into the growth plate region of Grg5 null mice. Like the growth defect, the growth plate and trabecular bone abnormality improved as the mice grew older. The growth plate defect was associated with reduced Indian hedgehog expression and signaling. We suggest that Grg5, a transcriptional coregulator, modulates the activities of transcription factors, such as Cbfa1/Runx2 in vivo to affect Ihh expression and the function of long bone growth plates.

  19. maLPA1-null mice as an endophenotype of anxious depression.

    PubMed

    Moreno-Fernández, R D; Pérez-Martín, M; Castilla-Ortega, E; Rosell Del Valle, C; García-Fernández, M I; Chun, J; Estivill-Torrús, G; Rodríguez de Fonseca, F; Santín, L J; Pedraza, C

    2017-04-04

    Anxious depression is a prevalent disease with devastating consequences and a poor prognosis. Nevertheless, the neurobiological mechanisms underlying this mood disorder remain poorly characterized. The LPA1 receptor is one of the six characterized G protein-coupled receptors (LPA1-6) through which lysophosphatidic acid acts as an intracellular signalling molecule. The loss of this receptor induces anxiety and several behavioural and neurobiological changes that have been strongly associated with depression. In this study, we sought to investigate the involvement of the LPA1 receptor in mood. We first examined hedonic and despair-like behaviours in wild-type and maLPA1 receptor null mice. Owing to the behavioural response exhibited by the maLPA1-null mice, the panic-like reaction was assessed. In addition, c-Fos expression was evaluated as a measure of the functional activity, followed by interregional correlation matrices to establish the brain map of functional activation. maLPA1-null mice exhibited anhedonia, agitation and increased stress reactivity, behaviours that are strongly associated with the psychopathological endophenotype of depression with anxiety features. Furthermore, the functional brain maps differed between the genotypes. The maLPA1-null mice showed increased limbic-system activation, similar to that observed in depressive patients. Antidepressant treatment induced behavioural improvements and functional brain normalisation. Finally, based on validity criteria, maLPA1-null mice are proposed as an animal model of anxious depression. Here, for we believe the first time, we have identified a possible relationship between the LPA1 receptor and anxious depression, shedding light on the unknown neurobiological basis of this subtype of depression and providing an opportunity to explore new therapeutic targets for the treatment of mood disorders, especially for the anxious subtype of depression.

  20. Inflammation enhances resection-induced intestinal adaptive growth in IL-10 null mice

    PubMed Central

    Speck, Karen E.; Garrison, Aaron P.; Rigby, Rachael J.; von Allmen, Doug C.; Lund, P. Kay; Helmrath, Michael A.

    2009-01-01

    Background Surgical resection of the ileum, cecum and proximal right colon (ICR) is common in the management of Crohn’s disease, yet little is known about the effect of active inflammation on the adaptive response following intestinal loss. We recently developed a surgical model of ICR in germ-free (GF) IL-10 null mice that develop small intestinal inflammation only when mice undergo conventionalization with normal fecal microflora (CONV) before surgical intervention. In this study, we examined the effects of post-surgical small bowel inflammation on adaptive growth after ICR. Methods 8-10 week old GF 129SvEv IL-10 null mice were allocated to GF or CONV groups. Non-operated GF and CONV mice provided baseline controls. Two weeks later GF and CONV mice were further allocated to ICR or sham operation. Small intestine and colon were harvested 7d after surgery for histological analysis. Results All mice within the gnotobiotic facility maintained GF status and did not develop small intestinal or colonic inflammation. CONV resulted in colitis in all groups, whereas small intestinal inflammation was only observed following ICR. Resection-induced small intestinal inflammation in CONV mice was associated with increases in proliferation, crypt depth and villus height when compared to GF mice after ICR. Resection-induced increases in crypt fission only occurred in CONV mice. Conclusion ICR-dependent small intestinal inflammation in CONV IL-10 null mice dramatically enhances early adaptive growth of the small intestine. Additional studies utilizing our model may provide clinical insight leading to optimal therapies in managing IBD patients after surgical resection. PMID:20074747

  1. Altered lipid and salt taste responsivity in ghrelin and GOAT null mice.

    PubMed

    Cai, Huan; Cong, Wei-Na; Daimon, Caitlin M; Wang, Rui; Tschöp, Matthias H; Sévigny, Jean; Martin, Bronwen; Maudsley, Stuart

    2013-01-01

    Taste perception plays an important role in regulating food preference, eating behavior and energy homeostasis. Taste perception is modulated by a variety of factors, including gastric hormones such as ghrelin. Ghrelin can regulate growth hormone release, food intake, adiposity, and energy metabolism. Octanoylation of ghrelin by ghrelin O-acyltransferase (GOAT) is a specific post-translational modification which is essential for many biological activities of ghrelin. Ghrelin and GOAT are both widely expressed in many organs including the gustatory system. In the current study, overall metabolic profiles were assessed in wild-type (WT), ghrelin knockout (ghrelin(-/-)), and GOAT knockout (GOAT(-/-)) mice. Ghrelin(-/-) mice exhibited decreased food intake, increased plasma triglycerides and increased ketone bodies compared to WT mice while demonstrating WT-like body weight, fat composition and glucose control. In contrast GOAT(-/-) mice exhibited reduced body weight, adiposity, resting glucose and insulin levels compared to WT mice. Brief access taste behavioral tests were performed to determine taste responsivity in WT, ghrelin(-/-) and GOAT(-/-) mice. Ghrelin and GOAT null mice possessed reduced lipid taste responsivity. Furthermore, we found that salty taste responsivity was attenuated in ghrelin(-/-) mice, yet potentiated in GOAT(-/-) mice compared to WT mice. Expression of the potential lipid taste regulators Cd36 and Gpr120 were reduced in the taste buds of ghrelin and GOAT null mice, while the salt-sensitive ENaC subunit was increased in GOAT(-/-) mice compared with WT mice. The altered expression of Cd36, Gpr120 and ENaC may be responsible for the altered lipid and salt taste perception in ghrelin(-/-) and GOAT(-/-) mice. The data presented in the current study potentially implicates ghrelin signaling activity in the modulation of both lipid and salt taste modalities.

  2. Altered Lipid and Salt Taste Responsivity in Ghrelin and GOAT Null Mice

    PubMed Central

    Daimon, Caitlin M.; Wang, Rui; Tschöp, Matthias H.; Sévigny, Jean; Martin, Bronwen; Maudsley, Stuart

    2013-01-01

    Taste perception plays an important role in regulating food preference, eating behavior and energy homeostasis. Taste perception is modulated by a variety of factors, including gastric hormones such as ghrelin. Ghrelin can regulate growth hormone release, food intake, adiposity, and energy metabolism. Octanoylation of ghrelin by ghrelin O-acyltransferase (GOAT) is a specific post-translational modification which is essential for many biological activities of ghrelin. Ghrelin and GOAT are both widely expressed in many organs including the gustatory system. In the current study, overall metabolic profiles were assessed in wild-type (WT), ghrelin knockout (ghrelin−/−), and GOAT knockout (GOAT−/−) mice. Ghrelin−/− mice exhibited decreased food intake, increased plasma triglycerides and increased ketone bodies compared to WT mice while demonstrating WT-like body weight, fat composition and glucose control. In contrast GOAT−/− mice exhibited reduced body weight, adiposity, resting glucose and insulin levels compared to WT mice. Brief access taste behavioral tests were performed to determine taste responsivity in WT, ghrelin−/− and GOAT−/− mice. Ghrelin and GOAT null mice possessed reduced lipid taste responsivity. Furthermore, we found that salty taste responsivity was attenuated in ghrelin−/− mice, yet potentiated in GOAT−/− mice compared to WT mice. Expression of the potential lipid taste regulators Cd36 and Gpr120 were reduced in the taste buds of ghrelin and GOAT null mice, while the salt-sensitive ENaC subunit was increased in GOAT−/− mice compared with WT mice. The altered expression of Cd36, Gpr120 and ENaC may be responsible for the altered lipid and salt taste perception in ghrelin−/− and GOAT−/− mice. The data presented in the current study potentially implicates ghrelin signaling activity in the modulation of both lipid and salt taste modalities. PMID:24124572

  3. ACE2 links amino acid malnutrition to microbial ecology and intestinal inflammation.

    PubMed

    Hashimoto, Tatsuo; Perlot, Thomas; Rehman, Ateequr; Trichereau, Jean; Ishiguro, Hiroaki; Paolino, Magdalena; Sigl, Verena; Hanada, Toshikatsu; Hanada, Reiko; Lipinski, Simone; Wild, Birgit; Camargo, Simone M R; Singer, Dustin; Richter, Andreas; Kuba, Keiji; Fukamizu, Akiyoshi; Schreiber, Stefan; Clevers, Hans; Verrey, Francois; Rosenstiel, Philip; Penninger, Josef M

    2012-07-25

    Malnutrition affects up to one billion people in the world and is a major cause of mortality. In many cases, malnutrition is associated with diarrhoea and intestinal inflammation, further contributing to morbidity and death. The mechanisms by which unbalanced dietary nutrients affect intestinal homeostasis are largely unknown. Here we report that deficiency in murine angiotensin I converting enzyme (peptidyl-dipeptidase A) 2 (Ace2), which encodes a key regulatory enzyme of the renin-angiotensin system (RAS), results in highly increased susceptibility to intestinal inflammation induced by epithelial damage. The RAS is known to be involved in acute lung failure, cardiovascular functions and SARS infections. Mechanistically, ACE2 has a RAS-independent function, regulating intestinal amino acid homeostasis, expression of antimicrobial peptides, and the ecology of the gut microbiome. Transplantation of the altered microbiota from Ace2 mutant mice into germ-free wild-type hosts was able to transmit the increased propensity to develop severe colitis. ACE2-dependent changes in epithelial immunity and the gut microbiota can be directly regulated by the dietary amino acid tryptophan. Our results identify ACE2 as a key regulator of dietary amino acid homeostasis, innate immunity, gut microbial ecology, and transmissible susceptibility to colitis. These results provide a molecular explanation for how amino acid malnutrition can cause intestinal inflammation and diarrhoea.

  4. Temporal and regional alterations in NMDA receptor expression in Mecp2-null mice.

    PubMed

    Blue, Mary E; Kaufmann, Walter E; Bressler, Joseph; Eyring, Charlotte; O'driscoll, Cliona; Naidu, Sakkubai; Johnston, Michael V

    2011-10-01

    Our previous postmortem study of girls with Rett Syndrome (RTT), a development disorder caused by MECP2 mutations, found increases in the density of N-Methyl-D-aspartate (NMDA) receptors in the prefrontal cortex of 2-8-year-old girls, whereas girls older than 10 years had reductions in NMDA receptors compared with age-matched controls (Blue et al., Ann Neurol 1999b;45:541-545). Using [(3)H]-CGP to label NMDA-type glutamate receptors in 2- and 7-week old wild-type (WT), Mecp2-null, and Mecp2-heterozygous (HET) mice (Bird model), we found that frontal areas of the brain also exhibited a bimodal pattern in NMDA expression, with increased densities of NMDA receptors in Mecp2-null mice at 2 weeks of age but decreased densities at 7 weeks of age. Visual cortex showed a similar pattern, while other cortical regions only exhibited changes in NMDA receptor densities at 2 weeks (retrosplenial granular) or 7 weeks (somatosensory). In thalamus of null mice, NMDA receptors were increased at 2 and 7 weeks. No significant differences in density were found between HET and WT mice at both ages. Western blots for NMDAR1 expression in frontal brain showed higher levels of expression in Mecp2-null mice at 2 weeks of age but not at 1 or 7 weeks of age. Our mouse data support the notion that deficient MeCP2 function is the primary cause of the NMDA receptor changes we observed in RTT. Furthermore, the findings of regional and temporal differences in NMDA expression illustrate the importance of age and brain region in evaluating different genotypes of mice.

  5. The sweeter side of ACE2: physiological evidence for a role in diabetes.

    PubMed

    Bindom, Sharell M; Lazartigues, Eric

    2009-04-29

    Diabetes mellitus is a growing problem in all parts of the world. Both clinical trials and animal models of type I and type II diabetes have shown that hyperactivity of angiotensin-II (Ang-II) signaling pathways contribute to the development of diabetes and diabetic complications. Of clinical relevance, blockade of the renin-angiotensin system prevents new-onset diabetes and reduces the risk of diabetic complications. Angiotensin-converting enzyme (ACE) 2 is a recently discovered mono-carboxypeptidase and the first homolog of ACE. It is thought to inhibit Ang-II signaling cascades mostly by cleaving Ang-II to generate Ang-(1-7), which effects oppose Ang-II and are mediated by the Mas receptor. The enzyme is present in the kidney, liver, adipose tissue and pancreas. Its expression is elevated in the endocrine pancreas in diabetes and in the early phase during diabetic nephropathy. ACE2 is hypothesized to act in a compensatory manner in both diabetes and diabetic nephropathy. Recently, we have shown the presence of the Mas receptor in the mouse pancreas and observed a reduction in Mas receptor immuno-reactivity as well as higher fasting blood glucose levels in ACE2 knockout mice, indicating that these mice may be a new model to study the role of ACE2 in diabetes. In this review we will examine the role of the renin-angiotensin system in the physiopathology and treatment of diabetes and highlight the potential benefits of the ACE2/Ang-(1-7)/Mas receptor axis, focusing on recent data about ACE2.

  6. Brain-targeted ACE2 overexpression attenuates neurogenic hypertension by inhibiting COX mediated inflammation

    PubMed Central

    Sriramula, Srinivas; Xia, Huijing; Xu, Ping; Lazartigues, Eric

    2014-01-01

    Overactivity of the renin angiotensin system (RAS), oxidative stress, and cyclooxygenases (COX) in the brain are implicated in the pathogenesis of hypertension. We previously reported that Angiotensin-Converting Enzyme 2 (ACE2) overexpression in the brain attenuates the development of DOCA-salt hypertension, a neurogenic hypertension model with enhanced brain RAS and sympathetic activity. To elucidate the mechanisms involved, we investigated whether oxidative stress, mitogen activated protein kinase signaling and cyclooxygenase (COX) activation in the brain are modulated by ACE2 in neurogenic hypertension. DOCA-salt hypertension significantly increased expression of Nox-2 (+61 ±5 %), Nox-4 (+50 ±13 %) and nitrotyrosine (+89 ±32 %) and reduced activity of the antioxidant enzymes, catalase (−29 ±4 %) and SOD (−31 ±7 %), indicating increased oxidative stress in the brain of non-transgenic mice. This increased oxidative stress was attenuated in transgenic mice overexpressing ACE2 in the brain. DOCA-salt-induced reduction of nNOS expression (−26 ±7 %) and phosphorylated eNOS/total eNOS (−30 ±3 %), and enhanced phosphorylation of Akt and ERK1/2 in the paraventricular nucleus (PVN), were reversed by ACE2 overexpression. In addition, ACE2 overexpression blunted the hypertension-mediated increase in gene and protein expression of COX-1 and COX-2 in the PVN. Furthermore, gene silencing of either COX-1 or COX-2 in the brain, reduced microglial activation and accompanied neuro-inflammation, ultimately attenuating DOCA-salt hypertension. Together, these data provide evidence that brain ACE2 overexpression reduces oxidative stress and COX-mediated neuro-inflammation, improves anti-oxidant and nitric oxide signaling, and thereby attenuates the development of neurogenic hypertension. PMID:25489058

  7. Altered astrocyte morphology and vascular development in dystrophin-Dp71-null mice.

    PubMed

    Giocanti-Auregan, Audrey; Vacca, Ophélie; Bénard, Romain; Cao, Sijia; Siqueiros, Lourdes; Montañez, Cecilia; Paques, Michel; Sahel, José-Alain; Sennlaub, Florian; Guillonneau, Xavier; Rendon, Alvaro; Tadayoni, Ramin

    2016-05-01

    Understanding retinal vascular development is crucial because many retinal vascular diseases such as diabetic retinopathy (in adults) or retinopathy of prematurity (in children) are among the leading causes of blindness. Given the localization of the protein Dp71 around the retinal vessels in adult mice and its role in maintaining retinal homeostasis, the aim of this study was to determine if Dp71 was involved in astrocyte and vascular development regulation. An experimental study in mouse retinas was conducted. Using a dual immunolabeling with antibodies to Dp71 and anti-GFAP for astrocytes on retinal sections and isolated astrocytes, it was found that Dp71 was expressed in wild-type (WT) mouse astrocytes from early developmental stages to adult stage. In Dp71-null mice, a reduction in GFAP-immunopositive astrocytes was observed as early as postnatal day 6 (P6) compared with WT mice. Using real-time PCR, it was showed that Dp71 mRNA was stable between P1 and P6, in parallel with post-natal vascular development. Regarding morphology in Dp71-null and WT mice, a significant decrease in overall astrocyte process number in Dp71-null retinas at P6 to adult age was found. Using fluorescence-conjugated isolectin Griffonia simplicifolia on whole mount retinas, subsequent delay of developing vascular network at the same age in Dp71-null mice was found. An evidence that the Dystrophin Dp71, a membrane-associated cytoskeletal protein and one of the smaller Duchenne muscular dystrophy gene products, regulates astrocyte morphology and density and is associated with subsequent normal blood vessel development was provided.

  8. Female ROMK null mice manifest more severe Bartter II phenotype on renal function and higher PGE2 production

    PubMed Central

    Yan, Qingshang; Yang, Xinbo; Cantone, Alessandra; Giebisch, Gerhard; Hebert, Steven; Wang, Tong

    2008-01-01

    ROMK null mice with a high survival rate and varying severity of hydronephrosis provide a good model to study type II Bartter syndrome pathophysiology (26). During the development of such a colony, we found that more male than female null mice survived, 58.7% vs. 33.3%. To investigate the possible mechanism of this difference, we compared the survival rates, renal functions, degree of hydronephrosis, as well as PGE2 and TXB2 production between male and female ROMK wild-type and null mice. We observed that female ROMK Bartter's mice exhibited lower GFR (0.37 vs. 0.54 ml·min−1·100 g BW−1, P < 0.05) and higher fractional Na+ excretion (0.66% vs. 0.48%, P < 0.05) than male Bartter's. No significant differences in acid-base parameters, urinary K+ excretion, and plasma electrolyte concentrations were observed between sexes. In addition, we assessed the liquid retention rate in the kidney to evaluate the extent of hydronephrosis and observed that 67% of male and 90% of female ROMK null mice were hydronephrotic mice. Urinary PGE2 excretion was higher in both sexes of ROMK null mice: 1.35 vs. 1.10 ng/24 h in males and 2.90 vs. 0.87 ng/24 h in females. TXB2 excretion was higher in female mice in both wild-type and ROMK null mice. The increments of urinary PGE2 and TXB2 were significantly higher in female null mice than males, 233.33% vs. 22.74% of PGE2 and 85.67% vs. 20.36% of TXB2. These data demonstrate a more severe Bartter phenotype in female ROMK null mice, and higher PGE2 and TXB2 production may be one of the mechanisms of this manifestation. PMID:18579648

  9. Amelogenesis imperfecta and other biomineralization defects in Fam20a and Fam20c null mice.

    PubMed

    Vogel, P; Hansen, G M; Read, R W; Vance, R B; Thiel, M; Liu, J; Wronski, T J; Smith, D D; Jeter-Jones, S; Brommage, R

    2012-11-01

    The FAM20 family of secreted proteins consists of three members (FAM20A, FAM20B, and FAM20C) recently linked to developmental disorders suggesting roles for FAM20 proteins in modulating biomineralization processes. The authors report here findings in knockout mice having null mutations affecting each of the three FAM20 proteins. Both Fam20a and Fam20c null mice survived to adulthood and showed biomineralization defects. Fam20b (-/-) embryos showed severe stunting and increased mortality at E13.5, although early lethality precluded detailed investigations. Physiologic calcification or biomineralization of extracellular matrices is a normal process in the development and functioning of various tissues (eg, bones and teeth). The lesions that developed in teeth, bones, or blood vessels after functional deletion of either Fam20a or Fam20c support a significant role for their encoded proteins in modulating biomineralization processes. Severe amelogenesis imperfecta (AI) was present in both Fam20a and Fam20c null mice. In addition, Fam20a (-/-) mice developed disseminated calcifications of muscular arteries and intrapulmonary calcifications, similar to those of fetuin-A deficient mice, although they were normocalcemic and normophosphatemic, with normal dentin and bone. Fam20a gene expression was detected in ameloblasts, odontoblasts, and the parathyroid gland, with local and systemic effects suggesting both local and/or systemic effects for FAM20A. In contrast, Fam20c (-/-) mice lacked ectopic calcifications but were severely hypophosphatemic and developed notable lesions in both dentin and bone to accompany the AI. The bone and dentin lesions, plus the marked hypophosphatemia and elevated serum alkaline phosphatase and FGF23 levels, are indicative of autosomal recessive hypophosphatemic rickets/osteomalacia in Fam20c (-/-) mice.

  10. Orexin Receptor Antagonism Improves Sleep and Reduces Seizures in Kcna1-null Mice

    PubMed Central

    Roundtree, Harrison M.; Simeone, Timothy A.; Johnson, Chaz; Matthews, Stephanie A.; Samson, Kaeli K.; Simeone, Kristina A.

    2016-01-01

    Study Objective: Comorbid sleep disorders occur in approximately one-third of people with epilepsy. Seizures and sleep disorders have an interdependent relationship where the occurrence of one can exacerbate the other. Orexin, a wake-promoting neuropeptide, is associated with sleep disorder symptoms. Here, we tested the hypothesis that orexin dysregulation plays a role in the comorbid sleep disorder symptoms in the Kcna1-null mouse model of temporal lobe epilepsy. Methods: Rest-activity was assessed using infrared beam actigraphy. Sleep architecture and seizures were assessed using continuous video-electroencephalography-electromyography recordings in Kcna1-null mice treated with vehicle or the dual orexin receptor antagonist, almorexant (100 mg/kg, intraperitoneally). Orexin levels in the lateral hypothalamus/perifornical region (LH/P) and hypothalamic pathology were assessed with immunohistochemistry and oxygen polarography. Results: Kcna1-null mice have increased latency to rapid eye movement (REM) sleep onset, sleep fragmentation, and number of wake epochs. The numbers of REM and non-REM (NREM) sleep epochs are significantly reduced in Kcna1-null mice. Severe seizures propagate to the wake-promoting LH/P where injury is apparent (indicated by astrogliosis, blood-brain barrier permeability, and impaired mitochondrial function). The number of orexin-positive neurons is increased in the LH/P compared to wild-type LH/P. Treatment with a dual orexin receptor antagonist significantly increases the number and duration of NREM sleep epochs and reduces the latency to REM sleep onset. Further, almorexant treatment reduces the incidence of severe seizures and overall seizure burden. Interestingly, we report a significant positive correlation between latency to REM onset and seizure burden in Kcna1-null mice. Conclusion: Dual orexin receptor antagonists may be an effective sleeping aid in epilepsy, and warrants further study on their somnogenic and ant-seizure effects in

  11. Glial dysfunction in parkin null mice: effects of aging.

    PubMed

    Solano, Rosa M; Casarejos, Maria J; Menéndez-Cuervo, Jamie; Rodriguez-Navarro, Jose A; García de Yébenes, Justo; Mena, Maria A

    2008-01-16

    Parkin mutations in humans produce parkinsonism whose pathogenesis is related to impaired protein degradation, increased free radicals, and abnormal neurotransmitter release. The role of glia in parkin deficiency is little known. We cultured midbrain glia from wild-type (WT) and parkin knock-out (PK-KO) mice. After 18-20 d in vitro, PK-KO glial cultures had less astrocytes, more microglia, reduced proliferation, and increased proapoptotic protein expression. PK-KO glia had greater levels of intracellular glutathione (GSH), increased mRNA expression of the GSH-synthesizing enzyme gamma-glutamylcysteine synthetase, and greater glutathione S-transferase and lower glutathione peroxidase activities than WT. The reverse happened in glia cultured in serum-free defined medium (EF12) or in old cultures. PK-KO glia was more susceptible than WT to transference to EF12 or neurotoxins (1-methyl-4-phenylpyridinium, blockers of GSH synthesis or catalase, inhibitors of extracellular signal-regulated kinase 1/2 and phosphatidylinositol 3 kinases), aging of the culture, or combination of these insults. PK-KO glia was less susceptible than WT to Fe2+ plus H2O2 and less responsive to protection by deferoxamine. Old WT glia increased the expression of heat shock protein 70, but PK-KO did not. Glia conditioned medium (GCM) from PK-KO was less neuroprotective and had lower levels of GSH than WT. GCM from WT increased the levels of dopamine markers in midbrain neuronal cultures transferred to EF12 more efficiently than GCM from PK-KO, and the difference was corrected by supplementation with GSH. PK-KO-GCM was a less powerful suppressor of apoptosis and microglia in neuronal cultures. Our data prove that abnormal glial function is critical in parkin mutations, and its role increases with aging.

  12. CRSBP-1/LYVE-1-null Mice Exhibit Identifiable Morphological and Functional Alterations of Lymphatic Capillary Vessels

    PubMed Central

    Huang, Shuan S.; Liu, I-Hua; Smith, Tracy; Shah, Maulik R.; Johnson, Frank E.; Huang, Jung S.

    2010-01-01

    CRSBP-1, a membrane glycoprotein, can mediate cell-surface retention of secreted growth factors containing CRS motifs such as PDGF-BB. CRSBP-1 has recently been found to be identical to LYVE-1, a specific marker for lymphatic capillary endothelial cells. The in vivo role of CRSBP-1/LYVE-1 is unknown. CRSBP-1-null mice are overtly normal and fertile but exhibit identifiable morphological and functional alterations of lymphatic capillary vessels in certain tissues, marked by the constitutively increased interstitial-lymphatic flow and lack of typical irregularly-shaped lumens. The CRSBP-1 ligands PDGF-BB and HA enhance interstitial-lymphatic flow in wild-type mice but not in CRSBP-1-null animals. PMID:17070806

  13. NOD/Shi-scid IL2rgamma(null) (NOG) mice more appropriate for humanized mouse models.

    PubMed

    Ito, M; Kobayashi, K; Nakahata, T

    2008-01-01

    "Humanized mice," in which various kinds of human cells and tissues can be engrafted and retain the same functions as in humans, are extremely useful because human diseases can be studied directly. Using the newly combined immunodeficient NOD-scid IL2rgamma(null) mice and Rag2(null) IL2rgamma(null) humanized mice, it has became possible to expand applications because various hematopoietic cells can be differentiated by human hematopoietic stem cell transplantation, and the human immune system can be reconstituted to some degree. This work has attracted attention worldwide, but the development and use of immunodeficient mice in Japan are not very well known or understood. This review describes the history and characteristics of the NOD/Shi-scid IL2rgamma(null) (NOG) and BALB/cA-Rag2(null) IL2rgamma(null) mice that were established in Japan, including our unpublished data from researchers who are currently using these mice. In addition, we also describe the potential development of new immunodeficient mice that can be used as humanized mice in the future.

  14. Neuronal Over-expression of ACE2 Protects Brain from Ischemia-induced Damage

    PubMed Central

    Chen, Ji; Zhao, Yuhui; Chen, Shuzhen; Wang, Jinju; Xiao, Xiang; Ma, Xiaotang; Penchikala, Madhuri; Xia, Huijing; Lazartigues, Eric; Zhao, Bin; Chen, Yanfang

    2014-01-01

    Angiotensin (Ang) II exaggerates cerebral injury in ischemic damage. Angiotensin-converting enzyme type 2 (ACE2) converts Ang II into Ang (1–7) and thus, may protect against the effects of Ang II. We hypothesized that neuronal ACE2 over-expression decreases ischemic stroke in mice with Ang II overproduction. Human renin and angiotensinogen double transgenic (RA) mice and RA mice with neuronal over-expression of ACE2 (SARA) were used for the study. The mean arterial pressure (MAP) was calculated from telemetry-recorded blood pressure (BP). SARA mice were infused peripherally with Norepinephrine to “clamp” the BP, or intracerebroventricularly-infused with a Mas receptor antagonist (A-779). Middle cerebral artery occlusion (MCAO) surgery was performed to induce permanent focal ischemic stroke. Cerebral blood flow (CBF) and neurological function were determined. Two days after surgery, brain samples were collected for various analyses. Results showed: 1) When compared to chronically hypertensive RA mice, SARA mice had lower basal MAP, less MCAO-induced infarct volume, and increased CBF, neurological function and cerebral microvascular density in the peri-infarct area; 2) These changes in SARA mice were not altered after MAP “clamping”, but partially reversed by brain infusion of A-779; 3) Ang (1–7)/Ang II ratio, angiogenic factors, endothelial nitric oxide synthase (eNOS) expression and nitric oxide production were increased, whereas, NADPH oxidase subunits and reactive oxygen species were decreased in the brain of SARA mice. ACE2 protects brain from ischemic injury via the regulation of NADPH oxidase/eNOS pathways by changing Ang (1–7)/Ang II ratio, independently of MAP changes. PMID:24440367

  15. Leucine supplementation via drinking water reduces atherosclerotic lesions in apoE null mice

    PubMed Central

    Zhao, Yang; Dai, Xiao-yan; Zhou, Zhou; Zhao, Ge-xin; Wang, Xian; Xu, Ming-jiang

    2016-01-01

    Aim: Recent evidence suggests that the essential amino acid leucine may be involved in systemic cholesterol metabolism. In this study, we investigated the effects of leucine supplementation on the development of atherosclerosis in apoE null mice. Methods: ApoE null mice were fed with chow supplemented with leucine (1.5% w/v) in drinking water for 8 week. Aortic atherosclerotic lesions were examined using Oil Red O staining. Plasma lipoprotein-cholesterol levels were measured with fast protein liquid chromatography. Hepatic gene expression was detected using real-time PCR and Western blot analyses. Results: Leucine supplementation resulted in 57.6% reduction of aortic atherosclerotic lesion area in apoE null mice, accompanied by 41.2% decrease of serum LDL-C levels and 40.2% increase of serum HDL-C levels. The body weight, food intake and blood glucose level were not affected by leucine supplementation. Furthermore, leucine supplementation increased the expression of Abcg5 and Abcg8 (that were involved in hepatic cholesterol efflux) by 1.28- and 0.86-fold, respectively, and significantly increased their protein levels. Leucine supplementation also increased the expression of Srebf1, Scd1 and Pgc1b (that were involved in hepatic triglyceride metabolism) by 3.73-, 1.35- and 1.71-fold, respectively. Consequently, leucine supplementation resulted in 51.77% reduction of liver cholesterol content and 2.2-fold increase of liver triglyceride content. Additionally, leucine supplementation did not affect the serum levels of IL-6, IFN-γ, TNF-α, IL-10 and IL-12, but markedly decreased the serum level of MCP-1. Conclusion: Leucine supplementation effectively attenuates atherosclerosis in apoE null mice by improving the plasma lipid profile and reducing systemic inflammation. PMID:26687933

  16. Galectin-3–null mice display defective neutrophil clearance during acute inflammation

    PubMed Central

    Wright, Rachael D; Souza, Patricia R.; Flak, Magdalena B.; Thedchanamoorthy, Prasheetha; Norling, Lucy V.; Cooper, Dianne

    2017-01-01

    Galectin-3 has been associated with a plethora of proinflammatory functions because of its ability, among others, to promote neutrophil activation and because of the reduction in neutrophil recruitment in models of infection in Gal-3-null mice. Conversely, it has also been linked to resolution of inflammation through its actions as an opsonin and its ability to promote efferocytosis of apoptotic neutrophils. Using a self-resolving model of peritonitis, we have addressed the modulation and role of Gal-3 in acute inflammation. We have shown that Gal-3 expression is increased in neutrophils that travel to the inflamed peritoneum and that cellular localization of this lectin is modulated during the course of the inflammatory response. Furthermore, neutrophil recruitment to the inflamed peritoneum is increased in Gal-3–null mice during the course of the response, and that correlates with reduced numbers of monocytes/macrophages in the cavities of those mice, as well as reduced apoptosis and efferocytosis of Gal-3–null neutrophils. These data indicate a role for endogenous Gal-3 in neutrophil clearance during acute inflammation. PMID:27733579

  17. Compensatory changes in CYP expression in three different toxicology mouse models: CAR-null, Cyp3a-null, and Cyp2b9/10/13-null mice

    PubMed Central

    Kumar, Ramiya; Mota, Linda C.; Litoff, Elizabeth J.; Rooney, John P.; Boswell, W. Tyler; Courter, Elliott; Henderson, Charles M.; Hernandez, Juan P.; Corton, J. Christopher; Moore, David D.

    2017-01-01

    Targeted mutant models are common in mechanistic toxicology experiments investigating the absorption, metabolism, distribution, or elimination (ADME) of chemicals from individuals. Key models include those for xenosensing transcription factors and cytochrome P450s (CYP). Here we investigated changes in transcript levels, protein expression, and steroid hydroxylation of several xenobiotic detoxifying CYPs in constitutive androstane receptor (CAR)-null and two CYP-null mouse models that have subfamily members regulated by CAR; the Cyp3a-null and a newly described Cyp2b9/10/13-null mouse model. Compensatory changes in CYP expression that occur in these models may also occur in polymorphic humans, or may complicate interpretation of ADME studies performed using these models. The loss of CAR causes significant changes in several CYPs probably due to loss of CAR-mediated constitutive regulation of these CYPs. Expression and activity changes include significant repression of Cyp2a and Cyp2b members with corresponding drops in 6α- and 16β-testosterone hydroxylase activity. Further, the ratio of 6α-/15α-hydroxylase activity, a biomarker of sexual dimorphism in the liver, indicates masculinization of female CAR-null mice, suggesting a role for CAR in the regulation of sexually dimorphic liver CYP profiles. The loss of Cyp3a causes fewer changes than CAR. Nevertheless, there are compensatory changes including gender-specific increases in Cyp2a and Cyp2b. Cyp2a and Cyp2b were down-regulated in CAR-null mice, suggesting activation of CAR and potentially PXR following loss of the Cyp3a members. However, the loss of Cyp2b causes few changes in hepatic CYP transcript levels and almost no significant compensatory changes in protein expression or activity with the possible exception of 6α-hydroxylase activity. This lack of a compensatory response in the Cyp2b9/10/13-null mice is probably due to low CYP2B hepatic expression, especially in male mice. Overall, compensatory and

  18. Otx1 null mutant mice show partial segregation of sensory epithelia comparable to lamprey ears

    NASA Technical Reports Server (NTRS)

    Fritzsch, B.; Signore, M.; Simeone, A.

    2001-01-01

    We investigated the development of inner ear innervation in Otx1 null mutants, which lack a horizontal canal, between embryonic day 12 (E12) and postnatal day 7 (P7) with DiI and immunostaining for acetylated tubulin. Comparable to control animals, horizontal crista-like fibers were found to cross over the utricle in Otx1 null mice. In mutants these fibers extend toward an area near the endolymphatic duct, not to a horizontal crista. Most Otx1 null mutants had a small patch of sensory hair cells at this position. Measurement of the area of the utricular macula suggested it to be enlarged in Otx1 null mutants. We suggest that parts of the horizontal canal crista remain incorporated in the utricular sensory epithelium in Otx1 null mutants. Other parts of the horizontal crista appear to be variably segregated to form the isolated patch of hair cells identifiable by the unique fiber trajectory as representing the horizontal canal crista. Comparison with lamprey ear innervation reveals similarities in the pattern of innervation with the dorsal macula, a sensory patch of unknown function. SEM data confirm that all foramina are less constricted in Otx1 null mutants. We propose that Otx1 is not directly involved in sensory hair cell formation of the horizontal canal but affects the segregation of the horizontal canal crista from the utricle. It also affects constriction of the two main foramina in the ear, but not their initial formation. Otx1 is thus causally related to horizontal canal morphogenesis as well as morphogenesis of these foramina.

  19. The role of neutrophils in corneal wound healing in HO-2 null mice.

    PubMed

    Marrazzo, Giuseppina; Bellner, Lars; Halilovic, Adna; Li Volti, Giovanni; Drago, Filippo; Dunn, Michael W; Schwartzman, Michal Laniado

    2011-01-01

    Our studies demonstrated that Heme oxygenase (HO), in particular, the constitutive HO-2, is critical for a self-resolving inflammatory and repair response in the cornea. Epithelial injury in HO-2 null mice leads to impaired wound closure and chronic inflammation in the cornea. This study was undertaken to examine the possible relationship between HO-2 and the recruitment of neutrophils following a corneal surface injury in wild type (WT) and HO-2 knockout (HO-2(-/-)) mice treated with Gr-1 monoclonal antibody to deplete peripheral neutrophils. Epithelial injury was performed by removing the entire corneal epithelium. Infiltration of inflammatory cell into the cornea in response to injury was higher in HO-2(-/-) than in WT. However, the rate of corneal wound closure following neutrophil depletion was markedly inhibited in both WT and HO-2(-/-) mice by 60% and 85%, respectively. Neutropenia induced HO-1 expression in WT but not in HO-2(-/-) mice. Moreover, endothelial cells lacking HO-2 expressed higher levels of the Midkine and VE-cadherin and displayed strong adhesion to neutrophils suggesting that perturbation in endothelial cell function caused by HO-2 depletion underlies the increased infiltration of neutrophils into the HO-2(-/-) cornea. Moreover, the fact that neutropenia worsened epithelial healing of the injured cornea in both WT and HO-2(-/-) mice suggest that cells other than neutrophils contribute to the exaggerated inflammation and impaired wound healing seen in the HO-2 null cornea.

  20. Stem cell expansion during carcinogenesis in stem cell-depleted conditional telomeric repeat factor 2 null mutant mice.

    PubMed

    Bojovic, B; Ho, H-Y; Wu, J; Crowe, D L

    2013-10-24

    To examine the role of telomeric repeat-binding factor 2 (TRF2) in epithelial tumorigenesis, we characterized conditional loss of TRF2 expression in the basal layer of mouse epidermis. These mice exhibit some characteristics of dyskeratosis congenita, a human stem cell depletion syndrome caused by telomere dysfunction. The epidermis in conditional TRF2 null mice exhibited DNA damage response and apoptosis, which correlated with stem cell depletion. The stem cell population in conditional TRF2 null epidermis exhibited shorter telomeres than those in control mice. Squamous cell carcinomas induced in conditional TRF2 null mice developed with increased latency and slower growth due to reduced numbers of proliferating cells as the result of increased apoptosis. TRF2 null epidermal stem cells were found in both primary and metastatic tumors. Despite the low-grade phenotype of the conditional TRF2 null primary tumors, the number of metastatic lesions was similar to control cancers. Basal cells from TRF2 null tumors demonstrated extreme telomere shortening and dramatically increased numbers of telomeric signals by fluorescence in situ hybridization due to increased genomic instability and aneuploidy in these cancers. DNA damage response signals were detected at telomeres in TRF2 null tumor cells from these mice. The increased genomic instability in these tumors correlated with eightfold expansion of the transformed stem cell population compared with that in control cancers. We concluded that genomic instability resulting from loss of TRF2 expression provides biological advantages to the cancer stem cell population.

  1. Altered somatosensory barrel cortex refinement in the developing brain of Mecp2-null mice.

    PubMed

    Moroto, M; Nishimura, A; Morimoto, M; Isoda, K; Morita, T; Yoshida, M; Morioka, S; Tozawa, T; Hasegawa, T; Chiyonobu, T; Yoshimoto, K; Hosoi, H

    2013-11-06

    Rett syndrome (RTT) is a neurodevelopmental disorder caused by mutations in the methyl-CpG binding protein 2 (MeCP2) gene. In previous studies, monoaminergic dysfunctions have been detected in patients with RTT and in a murine model of RTT, the Mecp2-null mouse. Therefore, the pathogenesis of RTT is thought to involve impairments in the monoaminergic systems. However, there have been limited data showing that the impairment of monoamines leads to early symptoms during development. We used histochemistry to study the somatosensory barrel cortex in the B6.129P2(C)-Mecp2(tm1.1Bird) mouse model of RTT. The barrel cortex is widely used to investigate neuronal development and its regulation by various neurotransmitters including 5-HT. 5-HT levels were measured by high performance liquid chromatography with electrochemical detection (HPLC/EC), and serotonin transporter (SERT) and 5-HT1B receptor mRNAs were measured in the somatosensory cortex, thalamus and striatum on postnatal days (P) 10, P20 and P40. Mecp2-null mice (Mecp2-/y) had significantly smaller barrel fields than age-matched wild-type controls (Mecp2+/y) on P10 and P40, but the topographic map was accurately formed. Levels of 5-HT, and SERT and 5-HT1B receptor mRNA expression in the somatosensory cortex did not differ significantly between the Mecp2-null and wild-type mice on P10. However, thalamic 5-HT was reduced in Mecp2-null mice. Our data indicate that a lack of MeCP2 may disturb the refinement of the barrel cortex in the early postnatal period. Our findings suggest that a decrease in thalamic 5-HT might be involved in this phenomenon.

  2. Pulmonary neuroendocrine cells, airway innervation, and smooth muscle are altered in Cftr null mice.

    PubMed

    Pan, Jie; Luk, Catherine; Kent, Geraldine; Cutz, Ernest; Yeger, Herman

    2006-09-01

    The amine- and peptide-producing pulmonary neuroendocrine cells (PNEC) are widely distributed within the airway mucosa of mammalian lung as solitary cells and innervated clusters, neuroepithelial bodies (NEB), which function as airway O2 sensors. These cells express Cftr and hence could play a role in the pathophysiology of cystic fibrosis (CF) lung disease. We performed confocal microscopy and morphometric analysis on lung sections from Cftr-/- (null), Cftr+/+, and Cftr+/- (control) mice at developmental stages E20, P5, P9, and P30 to determine the distribution, frequency, and innervation of PNEC/NEB, innervation and cell mass of airway smooth muscle, and neuromuscular junctions using synaptic vesicle protein 2, smooth muscle actin, and synaptophysin markers, respectively. The mean number of PNEC/NEB in Cftr-/- mice was significantly reduced compared with control mice at E20, whereas comparable or increased numbers were observed postnatally. NEB cells in Cftr null mice showed a significant reduction in intracorpuscular nerve endings compared with control mice, which is consistent with an intrinsic abnormality of the PNEC system. The airways of Cftr-/- mice showed reduced density (approximately 20-30%) of smooth muscle innervation, decreased mean airway smooth muscle mass (approximately 35%), and reduced density (approximately 20%) of nerve endings compared with control mice. We conclude that the airways of Cftr-/- mice exhibit heretofore unappreciated structural alterations affecting cellular and neural components of the PNEC system and airway smooth muscle and its innervation resulting in blunted O2 sensing and reduced airway tonus. Cftr could play a role in the development of the PNEC system, lung innervation, and airway smooth muscle.

  3. Modafinil more effectively induces wakefulness in orexin-null mice than in wild-type littermates.

    PubMed

    Willie, J T; Renthal, W; Chemelli, R M; Miller, M S; Scammell, T E; Yanagisawa, M; Sinton, C M

    2005-01-01

    Narcolepsy-cataplexy, a disorder of excessive sleepiness and abnormalities of rapid eye movement (REM) sleep, results from deficiency of the hypothalamic orexin (hypocretin) neuropeptides. Modafinil, an atypical wakefulness-promoting agent with an unknown mechanism of action, is used to treat hypersomnolence in these patients. Fos protein immunohistochemistry has previously demonstrated that orexin neurons are activated after modafinil administration, and it has been hypothesized that the wakefulness-promoting properties of modafinil might therefore be mediated by the neuropeptide. Here we tested this hypothesis by immunohistochemical, electroencephalographic, and behavioral methods using modafinil at doses of 0, 10, 30 and 100 mg/kg i.p. in orexin-/- mice and their wild-type littermates. We found that modafinil produced similar patterns of neuronal activation, as indicated by Fos immunohistochemistry, in both genotypes. Surprisingly, modafinil more effectively increased wakefulness time in orexin-/- mice than in the wild-type mice. This may reflect compensatory facilitation of components of central arousal in the absence of orexin in the null mice. In contrast, the compound did not suppress direct transitions from wakefulness to REM sleep, a sign of narcolepsy-cataplexy in mice. Spectral analysis of the electroencephalogram in awake orexin-/- mice under baseline conditions revealed reduced power in the theta; band frequencies (8-9 Hz), an index of alertness or attention during wakefulness in the rodent. Modafinil administration only partly compensated for this attention deficit in the orexin null mice. We conclude that the presence of orexin is not required for the wakefulness-prolonging action of modafinil, but orexin may mediate some of the alerting effects of the compound.

  4. Metabolic and Phenotypic Differences between Mice Producing a Werner Syndrome Helicase Mutant Protein and Wrn Null Mice

    PubMed Central

    Aumailley, Lucie; Garand, Chantal; Dubois, Marie Julie; Johnson, F. Brad; Marette, André; Lebel, Michel

    2015-01-01

    Werner syndrome (WS) is a premature aging disorder caused by mutations in a RecQ-family DNA helicase, WRN. Mice lacking part of the helicase domain of the WRN orthologue exhibit many phenotypic features of WS, including metabolic abnormalities and a shorter mean life span. In contrast, mice lacking the entire Wrn protein (i.e. Wrn null mice) do not exhibit a premature aging phenotype. In this study, we used a targeted mass spectrometry-based metabolomic approach to identify serum metabolites that are differentially altered in young Wrn helicase mutant and Wrn null mice. An antibody-based quantification of 43 serum cytokines and markers of cardiovascular disease risk complemented this study. We found that Wrn helicase mutants exhibited elevated and decreased levels, respectively, of the anti-inflammatory cytokine IL-10 and the pro-inflammatory cytokine IL-18. Wrn helicase mutants also exhibited an increase in serum hydroxyproline and plasminogen activator inhibitor-1, markers of extracellular matrix remodeling of the vascular system and inflammation in aging. We also observed an abnormal increase in the ratio of very long chain to short chain lysophosphatidylcholines in the Wrn helicase mutants underlying a peroxisome perturbation in these mice. Remarkably, the Wrn mutant helicase protein was mislocalized to the endoplasmic reticulum and the peroxisomal fractions in liver tissues. Additional analyses with mouse embryonic fibroblasts indicated a severe defect of the autophagy flux in cells derived from Wrn helicase mutants compared to wild type and Wrn null animals. These results indicate that the deleterious effects of the helicase-deficient Wrn protein are mediated by the dysfunction of several cellular organelles. PMID:26447695

  5. Enhanced endotoxin sensitivity in fps/fes-null mice with minimal defects in hematopoietic homeostasis.

    PubMed

    Zirngibl, Ralph A; Senis, Yotis; Greer, Peter A

    2002-04-01

    The fps/fes proto-oncogene encodes a cytoplasmic protein tyrosine kinase implicated in growth factor and cytokine receptor signaling and thought to be essential for the survival and terminal differentiation of myeloid progenitors. Fps/Fes-null mice were healthy and fertile, displayed slightly reduced numbers of bone marrow myeloid progenitors and circulating mature myeloid cells, and were more sensitive to lipopolysaccharide (LPS). These phenotypes were rescued using a fps/fes transgene. This confirmed that Fps/Fes is involved in, but not required for, myelopoiesis and that it plays a role in regulating the innate immune response. Bone marrow-derived Fps/Fes-null macrophages showed no defects in granulocyte-macrophage colony-stimulating factor-, interleukin 6 (IL-6)-, or IL-3-induced activation of signal transducer and activator of transcription 3 (Stat3) and Stat5A or LPS-induced degradation of I kappa B or activation of p38, Jnk, Erk, or Akt.

  6. Systemic Analysis of Atg5-Null Mice Rescued from Neonatal Lethality by Transgenic ATG5 Expression in Neurons.

    PubMed

    Yoshii, Saori R; Kuma, Akiko; Akashi, Takumi; Hara, Taichi; Yamamoto, Atsushi; Kurikawa, Yoshitaka; Itakura, Eisuke; Tsukamoto, Satoshi; Shitara, Hiroshi; Eishi, Yoshinobu; Mizushima, Noboru

    2016-10-10

    Autophagy is a cytoplasmic degradation system that is important for starvation adaptation and cellular quality control. Previously, we reported that Atg5-null mice are neonatal lethal; however, the exact cause of their death remains unknown. Here, we show that restoration of ATG5 in the brain is sufficient to rescue Atg5-null mice from neonatal lethality. This suggests that neuronal dysfunction, including suckling failure, is the primary cause of the death of Atg5-null neonates, which would further be accelerated by nutrient insufficiency due to a systemic failure in autophagy. The rescued Atg5-null mouse model, as a resource, allows us to investigate the physiological roles of autophagy in the whole body after the neonatal period. These rescued mice demonstrate previously unappreciated abnormalities such as hypogonadism and iron-deficiency anemia. These observations provide new insights into the physiological roles of the autophagy factor ATG5.

  7. Chronic Toxoplasma gondii in Nurr1-Null Heterozygous Mice Exacerbates Elevated Open Field Activity

    PubMed Central

    Eells, Jeffrey B.; Varela-Stokes, Andrea; Guo-Ross, Shirley X.; Kummari, Evangel; Smith, Holly M.; Cox, Erin; Lindsay, David S.

    2015-01-01

    Latent infection with Toxoplasma gondii is common in humans (approximately 30% of the global population) and is a significant risk factor for schizophrenia. Since prevalence of T. gondii infection is far greater than prevalence of schizophrenia (0.5-1%), genetic risk factors are likely also necessary to contribute to schizophrenia. To test this concept in an animal model, Nurr1-null heterozygous (+/-) mice and wild-type (+/+) mice were evaluate using an emergence test, activity in an open field and with a novel object, response to bobcat urine and prepulse inhibition of the acoustic startle response (PPI) prior to and 6 weeks after infection with T. gondii. In the emergence test, T. gondii infection significantly decreased the amount of time spent in the cylinder. Toxoplasma gondii infection significantly elevated open field activity in both +/+ and +/- mice but this increase was significantly exacerbated in +/- mice. T. gondii infection reduced PPI in male +/- mice but this was not statistically significant. Aversion to bobcat urine was abolished by T. gondii infection in +/+ mice. In female +/- mice, aversion to bobcat urine remained after T. gondii infection while the male +/- mice showed no aversion to bobcat urine. Antibody titers of infected mice were a critical variable associated with changes in open field activity, such that an inverted U shaped relationship existed between antibody titers and the percent change in open field activity with a significant increase in activity at low and medium antibody titers but no effect at high antibody titers. These data demonstrate that the Nurr1 +/- genotype predisposes mice to T. gondii-induced alterations in behaviors that involve dopamine neurotransmission and are associated with symptoms of schizophrenia. We propose that these alterations in murine behavior were due to further exacerbation of the altered dopamine neurotransmission in Nurr1 +/- mice. PMID:25855987

  8. Reactivation of autophagy by spermidine ameliorates the myopathic defects of collagen VI-null mice.

    PubMed

    Chrisam, Martina; Pirozzi, Marinella; Castagnaro, Silvia; Blaauw, Bert; Polishchuck, Roman; Cecconi, Francesco; Grumati, Paolo; Bonaldo, Paolo

    2015-01-01

    Autophagy is a self-degradative process responsible for the clearance of damaged or unnecessary cellular components. We have previously found that persistence of dysfunctional organelles due to autophagy failure is a key event in the pathogenesis of COL6/collagen VI-related myopathies, and have demonstrated that reactivation of a proper autophagic flux rescues the muscle defects of Col6a1-null (col6a1(-/-)) mice. Here we show that treatment with spermidine, a naturally occurring nontoxic autophagy inducer, is beneficial for col6a1(-/-) mice. Systemic administration of spermidine in col6a1(-/-) mice reactivated autophagy in a dose-dependent manner, leading to a concurrent amelioration of the histological and ultrastructural muscle defects. The beneficial effects of spermidine, together with its being easy to administer and the lack of overt side effects, open the field for the design of novel nutraceutical strategies for the treatment of muscle diseases characterized by autophagy impairment.

  9. Joint dysfunction and functional decline in middle age myostatin null mice.

    PubMed

    Guo, Wen; Miller, Andrew D; Pencina, Karol; Wong, Siu; Lee, Amanda; Yee, Michael; Toraldo, Gianluca; Jasuja, Ravi; Bhasin, Shalender

    2016-02-01

    Since its discovery as a potent inhibitor for muscle development, myostatin has been actively pursued as a drug target for age- and disease-related muscle loss. However, potential adverse effects of long-term myostatin deficiency have not been thoroughly investigated. We report herein that male myostatin null mice (mstn(-/-)), in spite of their greater muscle mass compared to wild-type (wt) mice, displayed more significant functional decline from young (3-6months) to middle age (12-15months) than age-matched wt mice, measured as gripping strength and treadmill endurance. Mstn(-/-) mice displayed markedly restricted ankle mobility and degenerative changes of the ankle joints, including disorganization of bone, tendon and peri-articular connective tissue, as well as synovial thickening with inflammatory cell infiltration. Messenger RNA expression of several pro-osteogenic genes was higher in the Achilles tendon-bone insertion in mstn(-/-) mice than wt mice, even at the neonatal age. At middle age, higher plasma concentrations of growth factors characteristic of excessive bone remodeling were found in mstn(-/-) mice than wt controls. These data collectively indicate that myostatin may play an important role in maintaining ankle and wrist joint health, possibly through negative regulation of the pro-osteogenic WNT/BMP pathway.

  10. Increased angiogenic response in aortic explants of collagen XVIII/endostatin-null mice.

    PubMed

    Li, Qing; Olsen, Bjorn R

    2004-08-01

    Endostatin, a proteolytic fragment of basement membrane-associated collagen XVIII, has been shown to be a potent angiogenesis inhibitor both in vivo and in vitro when given at high concentrations. The precise molecular mechanisms by which it functions and whether or not it plays a role in physiological regulation of angiogenesis are not clear. In mice with targeted null alleles of Col18a1, there appears to be no major abnormality in vascular patterns or capillary density in most organs. Furthermore, the growth of experimental tumors is not increased. However, a detailed analysis of induced angiogenesis in these mice has not been performed. Therefore, we compared the angiogenic responses induced by in vitro culture of aortic explants from collagen XVIII/endostatin-null mice (ko) to wild-type (wt) littermates. We found a twofold increase in microvessel outgrowth in explants from ko mice, relative to wt explants. This increased angiogenesis was reduced to the wt level by the addition of low levels (0.1 microg/ml) of recombinant mouse or human endostatin during the culture period. To address cellular/molecular mechanisms underlying this difference in angiogenic response between ko and wt mice, we isolated endothelial cells from both strains and compared their biological behavior. Proliferation assays showed no difference between the two types of endothelial cells. In contrast, adhesion assays showed a striking difference in their ability to adhere to fibronectin suggesting that collagen XVIII/endostatin may regulate interactions between endothelial cells and underlying basement membrane-associated components, including fibronectin, such that in the absence of collagen XVIII/endostatin, endothelial cells are more adhesive to fibronectin. In the aortic explant assay, characterized by dynamic processes of microvessel elongation and regression, this may result in stabilization of newly formed vessels, reduced regression, and a net increase in microvessel outgrowth in

  11. Epithelial defect in prostates of Stat5a-null mice.

    PubMed

    Nevalainen, M T; Ahonen, T J; Yamashita, H; Chandrashekar, V; Bartke, A; Grimley, P M; Robinson, G W; Hennighausen, L; Rui, H

    2000-07-01

    The transcription factor Stat5a critically mediates prolactin (PRL)-induced mammary gland development and lactogenesis. PRL also stimulates growth and differentiation of prostate tissue. Specifically, hyperprolactinemia gives rise to prostate hyperplasia, and prostate size is reduced in PRL-deficient mice. We therefore investigated the importance of Stat5a for prostate development and function by examining Stat5a-null mice. The absence of Stat5a in mice was associated with a distinct prostate morphology characterized by an increased prevalence of local disorganization within acinar epithelium of ventral prostates. Affected acini were typically filled with desquamated, granular epithelial cells that had become embedded in dense, coagulated secretory material. These features were reminiscent of acinar cyst formation and degeneration frequently observed in human benign prostate hyperplasia, however, cystic changes in prostate acini of Stat5a-deficient mice were not associated with increased prostate size or morphologic hallmarks of epithelial hyperplasia. Instead, immunohistochemistry of the prostate-specific secretory marker, probasin, suggested that hypersecretory function of the epithelium could underlie local congestion and cyst formation in prostates of Stat5a-null mice. Serum testosterone and PRL levels were normal in Stat5a knockout mice, but prostate PRL receptor expression was reduced as determined by immunohistochemistry. Expression levels or activation states of other PRL signal transduction proteins, including Stat5b, Stat3, Stat1, ERK1, and ERK2 were not altered. The present study offers the first evidence for a direct role of Stat5a in the maintenance of normal tissue architecture and function of the mouse prostate.

  12. Uncovering Novel Reproductive Defects in Neurokinin B Receptor Null Mice: Closing the Gap Between Mice and Men

    PubMed Central

    Yang, Jasmine J.; Caligioni, Claudia S.; Chan, Yee-Ming

    2012-01-01

    Patients bearing mutations in TAC3 and TACR3 (which encode neurokinin B and its receptor, respectively) have sexual infantilism and infertility due to GnRH deficiency. In contrast, Tacr3−/− mice have previously been reported to be fertile. Because of this apparent phenotypic discordance between mice and men bearing disabling mutations in Tacr3/TACR3, Tacr3 null mice were phenotyped with close attention to pubertal development, estrous cyclicity, and fertility. Tacr3−/− mice demonstrated normal timing of preputial separation and day of first estrus, markers of sexual maturation. However, at postnatal d 60, Tacr3−/− males had significantly smaller testes and lower FSH levels than their wild-type littermates. Tacr3−/− females had lower uterine weights and abnormal estrous cyclicity. Approximately half of Tacr3−/− females had no detectable corpora lutea on ovarian histology at postnatal d 60. Despite this apparent ovulatory defect, all Tacr3−/− females achieved fertility when mated. However, Tacr3−/− females were subfertile, having both reduced numbers of litters and pups per litter. The subfertility of these animals was not due to a primary ovarian defect, because they demonstrated a robust response to exogenous gonadotropins. Thus, although capable of fertility, Tacr3-deficient mice have central reproductive defects. The remarkable ability of acyclic female Tacr3 null mice to achieve fertility is reminiscent of the reversal of hypogonadotropic hypogonadism seen in a high proportion of human patients bearing mutations in TACR3. Tacr3 mice are a useful model to examine the mechanisms by which neurokinin B signaling modulates GnRH release. PMID:22253416

  13. Altered ion-responsive gene expression in Mmp20 null mice.

    PubMed

    Tye, C E; Sharma, R; Smith, C E; Bartlett, J D

    2010-12-01

    During enamel maturation, hydroxyapatite crystallites expand in volume, releasing protons that acidify the developing enamel. This acidity is neutralized by the buffering activity of carbonic anhydrases and ion transporters. Less hydroxyapatite forms in matrix metalloproteinase-20 null (Mmp20(-/-)) mouse incisors, because enamel thickness is reduced by approximately 50%. We therefore asked if ion regulation was altered in Mmp20(-/-) mouse enamel. Staining of wild-type and Mmp20(-/-) incisors with pH indicators demonstrated that wild-type mice had pronounced changes in enamel pH as development progressed. These pH changes were greatly attenuated in Mmp20(-/-) mice. Expression of 4 ion-regulatory genes (Atp2b4, Slc4a2, Car6, Cftr) was significantly decreased in enamel organs from Mmp20(-/-) mice. Notably, expression of secreted carbonic anhydrase (Car6) was reduced to almost undetectable levels in the null enamel organ. In contrast, Odam and Klk4 expression was unaffected. We concluded that a feedback mechanism regulates ion-responsive gene expression during enamel development.

  14. Seizure phenotypes, periodicity, and sleep-wake pattern of seizures in Kcna-1 null mice.

    PubMed

    Wright, Samantha; Wallace, Eli; Hwang, Youngdeok; Maganti, Rama

    2016-02-01

    This study was undertaken to describe seizure phenotypes, natural progression, sleep-wake patterns, as well as periodicity of seizures in Kcna-1 null mutant mice. These mice were implanted with epidural electroencephalography (EEG) and electromyography (EMG) electrodes, and simultaneous video-EEG recordings were obtained while animals were individually housed under either diurnal (LD) condition or constant darkness (DD) over ten days of recording. The video-EEG data were analyzed to identify electrographic and behavioral phenotypes and natural progression and to examine the periodicity of seizures. Sleep-wake patterns were analyzed to understand the distribution and onset of seizures across the sleep-wake cycle. Four electrographically and behaviorally distinct seizure types were observed. Regardless of lighting condition that animals were housed in, Kcna-1 null mice initially expressed only a few of the most severe seizure types that progressively increased in frequency and decreased in seizure severity. In addition, a circadian periodicity was noted, with seizures peaking in the first 12h of the Zeitgeber time (ZT) cycle, regardless of lighting conditions. Interestingly, seizure onset differed between lighting conditions where more seizures arose out of sleep in LD conditions, whereas under DD conditions, the majority occurred out of the wakeful state. We suggest that this model be used to understand the circadian pattern of seizures as well as the pathophysiological implications of sleep and circadian disturbances in limbic epilepsies.

  15. Reduced excitatory neurotransmission and mild autism-relevant phenotypes in adolescent Shank3 null mutant mice.

    PubMed

    Yang, Mu; Bozdagi, Ozlem; Scattoni, Maria Luisa; Wöhr, Markus; Roullet, Florence I; Katz, Adam M; Abrams, Danielle N; Kalikhman, David; Simon, Harrison; Woldeyohannes, Leuk; Zhang, James Y; Harris, Mark J; Saxena, Roheeni; Silverman, Jill L; Buxbaum, Joseph D; Crawley, Jacqueline N

    2012-05-09

    Mutations in the synaptic scaffolding protein gene SHANK3 are strongly implicated in autism and Phelan-McDermid 22q13 deletion syndrome. The precise location of the mutation within the Shank3 gene is key to its phenotypic outcomes. Here, we report the physiological and behavioral consequences of null and heterozygous mutations in the ankyrin repeat domain in Shank3 mice. Both homozygous and heterozygous mice showed reduced glutamatergic transmission and long-term potentiation in the hippocampus with more severe deficits detected in the homozygous mice. Three independent cohorts were evaluated for magnitude and replicability of behavioral endophenotypes relevant to autism and Phelan-McDermid syndrome. Mild social impairments were detected, primarily in juveniles during reciprocal interactions, while all genotypes displayed normal adult sociability on the three-chambered task. Impaired novel object recognition and rotarod performance were consistent across cohorts of null mutants. Repetitive self-grooming, reduced ultrasonic vocalizations, and deficits in reversal of water maze learning were detected only in some cohorts, emphasizing the importance of replication analyses. These results demonstrate the exquisite specificity of deletions in discrete domains within the Shank3 gene in determining severity of symptoms.

  16. Generation of mice with a conditional null Fraser syndrome 1 (Fras1) allele.

    PubMed

    Pitera, Jolanta E; Turmaine, Mark; Woolf, Adrian S; Scambler, Peter J

    2012-12-01

    Fraser syndrome (FS) is an autosomal recessive disease characterized by skin lesions and kidney and upper airway malformations. Fraser syndrome 1 (FRAS1) is an extracellular matrix protein, and FRAS1 homozygous mutations occur in some FS individuals. FRAS1 is expressed at the epithelial-mesenchymal interface in embryonic skin and kidney. blebbed mice have a null Fras1 mutation and phenocopy human FS. Like humans with FS, they exhibit a high fetal and neonatal mortality, precluding studies of FRAS1 functions in later life. We generated conditional Fras1 null allele mice. Cre-mediated generalized deletion of this allele generated embryonic skin blisters and renal agenesis characteristic of blebbed mice and human FS. Targeted deletion of Fras1 in kidney podocytes circumvented skin blistering, renal agenesis, and early death. FRAS1 expression was downregulated in maturing glomeruli which then became sclerotic. The data are consistent with the hypothesis that locally produced FRAS1 has roles in glomerular maturation and integrity. This conditional allele will facilitate study of possible role for FRAS1 in other tissues such as the skin.

  17. Amphetamine-elicited striatal Fos expression is attenuated in neurotensin null mutant mice.

    PubMed

    Fadel, Jim; Dobner, Paul R; Deutch, Ariel Y

    2006-07-10

    Neurotensin (NT) has been suggested to interact with dopamine systems in different forebrain sites to exert both antipsychotic- and psychostimulant-like effects. We previously found that genetic or pharmacological manipulations that disrupt endogenous NT signaling attenuate antipsychotic drug-induced Fos expression in the dorsolateral and central striatum but not other striatal regions. To assess the role of NT in psychostimulant responses, we examined the ability of d-amphetamine (AMP) to induce Fos in wild-type and NT null mutant mice. AMP-elicited Fos expression was significantly attenuated in the medial striatum of NT null mutant mice, but was unaffected in other striatal territories. Similar results were obtained in rats and mice pretreated with the high affinity neurotensin receptor (NTR1) antagonist SR 48692. The effect of the NTR1 antagonist was particularly apparent in the striatal patch (striosome) compartment, as defined by mu-opioid receptor immunoreactivity. These data suggest that NT is required for the full activation by AMP of medial striatal neurons.

  18. The Lack of Thrombospondin-1 (TSP1) Dictates the Course of Wound Healing in Double-TSP1/TSP2-Null Mice

    PubMed Central

    Agah, Azin; Kyriakides, Themis R.; Lawler, Jack; Bornstein, Paul

    2002-01-01

    Thrombospondin (TSP) 1 and 2, share the same overall structure and interact with a number of the same cell-surface receptors. In an attempt to elucidate their biological roles more clearly, we generated double-TSP1/TSP2-null animals and compared their phenotype to those of TSP1- and TSP2-null mice. Double-null mice exhibited an apparent phenotype that primarily represented the sum of the abnormalities observed in the single-null mice. However, surprisingly, the wound-healing response in double-null mice resembled that in TSP1-null animals and differed from that in TSP2-nulls. Thus, although the excisional wounds of TSP2-null mice are characterized by increased neovascularization and heal at an accelerated rate, TSP1-null and double-null animals demonstrated delayed healing, as indicated by the prolonged persistence of inflammation and delayed scab loss. Immunohistochemical analysis showed that, similar to TSP1-null mice, the granulation tissue of double-null mice was not excessively vascularized. Furthermore as in TSP1-nulls, decreases in macrophage recruitment and in the levels of monocyte chemoattractant protein-1 indicated that the inflammatory phase of the wound-healing response was impaired in double-null mice. Our data demonstrate that the consequences of a lack of TSP1 predominate in the response of double-null mice, and dictate the course of wound healing. These findings reflect distinct temporal and spatial expressions of TSP1 and TSP2 in the healing wound. PMID:12213711

  19. ACE2 and vasoactive peptides: novel players in cardiovascular/renal remodeling and hypertension.

    PubMed

    Mendoza-Torres, Evelyn; Oyarzún, Alejandra; Mondaca-Ruff, David; Azocar, Andrés; Castro, Pablo F; Jalil, Jorge E; Chiong, Mario; Lavandero, Sergio; Ocaranza, María Paz

    2015-08-01

    The renin-angiotensin system (RAS) is a key component of cardiovascular physiology and homeostasis due to its influence on the regulation of electrolyte balance, blood pressure, vascular tone and cardiovascular remodeling. Deregulation of this system contributes significantly to the pathophysiology of cardiovascular and renal diseases. Numerous studies have generated new perspectives about a noncanonical and protective RAS pathway that counteracts the proliferative and hypertensive effects of the classical angiotensin-converting enzyme (ACE)/angiotensin (Ang) II/angiotensin type 1 receptor (AT1R) axis. The key components of this pathway are ACE2 and its products, Ang-(1-7) and Ang-(1-9). These two vasoactive peptides act through the Mas receptor (MasR) and AT2R, respectively. The ACE2/Ang-(1-7)/MasR and ACE2/Ang-(1-9)/AT2R axes have opposite effects to those of the ACE/Ang II/AT1R axis, such as decreased proliferation and cardiovascular remodeling, increased production of nitric oxide and vasodilation. A novel peptide from the noncanonical pathway, alamandine, was recently identified in rats, mice and humans. This heptapeptide is generated by catalytic action of ACE2 on Ang A or through a decarboxylation reaction on Ang-(1-7). Alamandine produces the same effects as Ang-(1-7), such as vasodilation and prevention of fibrosis, by interacting with Mas-related GPCR, member D (MrgD). In this article, we review the key roles of ACE2 and the vasoactive peptides Ang-(1-7), Ang-(1-9) and alamandine as counter-regulators of the ACE-Ang II axis as well as the biological properties that allow them to regulate blood pressure and cardiovascular and renal remodeling.

  20. Fibroblast Growth Factor 21-Null Mice Do Not Exhibit an Impaired Response to Fasting

    PubMed Central

    Antonellis, Patrick Joseph; Hayes, Meghan Patricia; Adams, Andrew Charles

    2016-01-01

    Fibroblast growth factor 21 (FGF21) is a pleotropic metabolic regulator, expression of which is elevated during fasting. To this end, the precise role played by FGF21 in the biology of fasting has been the subject of several recent studies, which have demonstrated contributions to the regulation of both lipid and carbohydrate metabolism. In the present study, we compared wild-type (WT) and FGF21-null (FGF21KO) mice, demonstrating that, despite the significant induction of FGF21 during fasting in the WT animals, our strain of FGF21-null mice exhibits only limited impairments in their adaptation to nutrient deprivation. Specifically, fasted FGF21KO mice display a mild attenuation of gluconeogenic transcriptional induction in the liver accompanied by partially blunted glucose production in response to a pyruvate challenge. Furthermore, FGF21KO mice displayed only minor impairments in lipid metabolism in the fasted state, limited to accumulation of hepatic triglycerides and a reduction in expression of genes associated with fatty acid oxidation. To address the possibility of compensation to germline deletion of FGF21, we further interrogated the role of endogenous FGF21 via acute pharmacological blockade of FGF21 signaling. At the transcriptional level, we show that FGF21 signaling is required for full induction of gluconeogenic and oxidative genes in the liver. However, corroborating our findings in FGF21KO mice, pharmacological blockade of the FGF21 axis did not profoundly disrupt the physiological response to fasting. Taken as a whole, these data demonstrate that, while FGF21 is partially required for appropriate gene expression during the fed to fasted transition, its absence does not significantly impact the downstream physiology of the fasted state. PMID:27445980

  1. ACE2 and Microbiota: Emerging Targets for Cardiopulmonary Disease Therapy

    PubMed Central

    Cole-Jeffrey, Colleen T; Liu, Meng; Katovich, Michael J; Raizada, Mohan K; Shenoy, Vinayak

    2015-01-01

    The health of the cardiovascular and pulmonary systems is inextricably linked to the renin-angiotensin system (RAS). Physiologically speaking, a balance between the vasodeleterious (ACE/Ang II/AT1R) and vasoprotective (ACE2/Ang-(1–7)/MasR) components of the RAS is critical for cardiopulmonary homeostasis. Upregulation of the ACE/Ang II/AT1R axis shifts the system toward vasoconstriction, proliferation, hypertrophy, inflammation, and fibrosis, all factors that contribute to the development and progression of cardiopulmonary diseases. Conversely, stimulation of the vasoprotective ACE2/Ang-(1–7)/MasR axis produces a counter-regulatory response that promotes cardiovascular health. Current research is investigating novel strategies to augment actions of the vasoprotective RAS components, particularly ACE2, in order to treat various pathologies. While multiple approaches to increase the activity of ACE2 have displayed beneficial effects against experimental disease models, the mechanisms behind its protective actions remain incompletely understood. Recent work demonstrating a non-catalytic role for ACE2 in amino acid transport in the gut has led us to speculate that the therapeutic effects of ACE2 can be mediated, in part, by its actions on the gastrointestinal tract and/or gut microbiome. This is consistent with emerging data which suggests that dysbiosis of the gut and lung microbiomes is associated with cardiopulmonary disease. This review highlights new developments in the protective actions of ACE2 against cardiopulmonary disorders, discusses innovative approaches to targeting ACE2 for therapy, and explores an evolving role for gut and lung microbiota in cardiopulmonary health. PMID:26322922

  2. ACE2 and Microbiota: Emerging Targets for Cardiopulmonary Disease Therapy.

    PubMed

    Cole-Jeffrey, Colleen T; Liu, Meng; Katovich, Michael J; Raizada, Mohan K; Shenoy, Vinayak

    2015-12-01

    The health of the cardiovascular and pulmonary systems is inextricably linked to the renin-angiotensin system (RAS). Physiologically speaking, a balance between the vasodeleterious (Angiotensin-converting enzyme [ACE]/Angiotensin II [Ang II]/Ang II type 1 receptor [AT1R]) and vasoprotective (Angiotensin-converting enzyme 2 [ACE2]/Angiotensin-(1-7) [Ang-(1-7)]/Mas receptor [MasR]) components of the RAS is critical for cardiopulmonary homeostasis. Upregulation of the ACE/Ang II/AT1R axis shifts the system toward vasoconstriction, proliferation, hypertrophy, inflammation, and fibrosis, all factors that contribute to the development and progression of cardiopulmonary diseases. Conversely, stimulation of the vasoprotective ACE2/Ang-(1-7)/MasR axis produces a counter-regulatory response that promotes cardiovascular health. Current research is investigating novel strategies to augment actions of the vasoprotective RAS components, particularly ACE2, in order to treat various pathologies. Although multiple approaches to increase the activity of ACE2 have displayed beneficial effects against experimental disease models, the mechanisms behind its protective actions remain incompletely understood. Recent work demonstrating a non-catalytic role for ACE2 in amino acid transport in the gut has led us to speculate that the therapeutic effects of ACE2 can be mediated, in part, by its actions on the gastrointestinal tract and/or gut microbiome. This is consistent with emerging data which suggest that dysbiosis of the gut and lung microbiomes is associated with cardiopulmonary disease. This review highlights new developments in the protective actions of ACE2 against cardiopulmonary disorders, discusses innovative approaches to targeting ACE2 for therapy, and explores an evolving role for gut and lung microbiota in cardiopulmonary health.

  3. Behavioral Disturbances in Estrogen-Related Receptor alpha-Null Mice

    PubMed Central

    Cui, Huxing; Lu, Yuan; Khan, Michael Z.; Anderson, Rachel M.; McDaniel, Latisha; Wilson, Hannah E.; Yin, Terry C.; Radley, Jason J.; Pieper, Andrew A.; Lutter, Michael

    2015-01-01

    SUMMARY Eating disorders, such as anorexia nervosa and bulimia nervosa, are common and severe mental illnesses of unknown etiology. Recently, we identified a rare missense mutation in the transcription factor estrogen-related receptor alpha (ESRRA) that is associated with the development of eating disorders. However, little is known about ESRRA function in the brain. Here, we report that Esrra is expressed in the mouse brain and demonstrate that Esrra levels are regulated by energy reserves. Esrra-null female mice display a reduced operant response to a high-fat diet, compulsivity/behavioral rigidity, and social deficits. Selective Esrra knockdown in the prefrontal and orbitofrontal cortices of adult female mice recapitulates reduced operant response and increased compulsivity, respectively. These results indicate that Esrra deficiency in the mouse brain impairs behavioral responses in multiple functional domains. PMID:25865889

  4. Altered Arachidonate Distribution in Macrophages from Caveolin-1 Null Mice Leading to Reduced Eicosanoid Synthesis*

    PubMed Central

    Astudillo, Alma M.; Pérez-Chacón, Gema; Meana, Clara; Balgoma, David; Pol, Albert; del Pozo, Miguel A.; Balboa, María A.; Balsinde, Jesús

    2011-01-01

    In this work we have studied the effect of caveolin-1 deficiency on the mechanisms that regulate free arachidonic acid (AA) availability. The results presented here demonstrate that macrophages from caveolin-1-deficient mice exhibit elevated fatty acid incorporation and remodeling and a constitutively increased CoA-independent transacylase activity. Mass spectrometry-based lipidomic analyses reveal stable alterations in the profile of AA distribution among phospholipids, manifested by reduced levels of AA in choline glycerophospholipids but elevated levels in ethanolamine glycerophospholipids and phosphatidylinositol. Furthermore, macrophages from caveolin-1 null mice show decreased AA mobilization and prostaglandin E2 and LTB4 production upon cell stimulation. Collectively, these results provide insight into the role of caveolin-1 in AA homeostasis and suggest an important role for this protein in the eicosanoid biosynthetic response. PMID:21852231

  5. Autism phenotypes in ZnT3 null mice: Involvement of zinc dyshomeostasis, MMP-9 activation and BDNF upregulation.

    PubMed

    Yoo, Min Heui; Kim, Tae-Youn; Yoon, Young Hee; Koh, Jae-Young

    2016-06-29

    To investigate the role of synaptic zinc in the ASD pathogenesis, we examined zinc transporter 3 (ZnT3) null mice. At 4-5 weeks of age, male but not female ZnT3 null mice exhibited autistic-like behaviors. Cortical volume and neurite density were significantly greater in male ZnT3 null mice than in WT mice. In male ZnT3 null mice, consistent with enhanced neurotrophic stimuli, the level of BDNF as well as activity of MMP-9 was increased. Consistent with known roles for MMPs in BDNF upregulation, 2.5-week treatment with minocycline, an MMP inhibitor, significantly attenuated BDNF levels as well as megalencephaly and autistic-like behaviors. Although the ZnT3 null state removed synaptic zinc, it rather increased free zinc in the cytosol of brain cells, which appeared to increase MMP-9 activity and BDNF levels. The present results suggest that zinc dyshomeostasis during the critical period of brain development may be a possible contributing mechanism for ASD.

  6. Autism phenotypes in ZnT3 null mice: Involvement of zinc dyshomeostasis, MMP-9 activation and BDNF upregulation

    PubMed Central

    Yoo, Min Heui; Kim, Tae-Youn; Yoon, Young Hee; Koh, Jae-Young

    2016-01-01

    To investigate the role of synaptic zinc in the ASD pathogenesis, we examined zinc transporter 3 (ZnT3) null mice. At 4–5 weeks of age, male but not female ZnT3 null mice exhibited autistic-like behaviors. Cortical volume and neurite density were significantly greater in male ZnT3 null mice than in WT mice. In male ZnT3 null mice, consistent with enhanced neurotrophic stimuli, the level of BDNF as well as activity of MMP-9 was increased. Consistent with known roles for MMPs in BDNF upregulation, 2.5-week treatment with minocycline, an MMP inhibitor, significantly attenuated BDNF levels as well as megalencephaly and autistic-like behaviors. Although the ZnT3 null state removed synaptic zinc, it rather increased free zinc in the cytosol of brain cells, which appeared to increase MMP-9 activity and BDNF levels. The present results suggest that zinc dyshomeostasis during the critical period of brain development may be a possible contributing mechanism for ASD. PMID:27352957

  7. Transcriptional Fingerprint of Hypomyelination in Zfp191null and Shiverer (Mbpshi) Mice.

    PubMed

    Aaker, Joshua D; Elbaz, Benayahu; Wu, Yuwen; Looney, Timothy J; Zhang, Li; Lahn, Bruce T; Popko, Brian

    2016-10-01

    The transcriptional program that controls oligodendrocyte maturation and central nervous system (CNS) myelination has not been fully characterized. In this study, we use high-throughput RNA sequencing to analyze how the loss of a key transcription factor, zinc finger protein 191 (ZFP191), results in oligodendrocyte development abnormalities and CNS hypomyelination. Using a previously described mutant mouse that is deficient in ZFP191 protein expression (Zfp191(null)), we demonstrate that key transcripts are reduced in the whole brain as well as within oligodendrocyte lineage cells cultured in vitro To determine whether the loss of myelin seen in Zfp191(null) mice contributes indirectly to these perturbations, we also examined the transcriptome of a well-characterized mouse model of hypomyelination, in which the myelin structural protein myelin basic protein (MBP) is deficient. Interestingly, Mbp(shi) (shiverer) mice had far fewer transcripts perturbed with the loss of myelin alone. This study demonstrates that the loss of ZFP191 disrupts expression of genes involved in oligodendrocyte maturation and myelination, largely independent from the loss of myelin. Nevertheless, hypomyelination in both mouse mutants results in the perturbation of lipid synthesis pathways, suggesting that oligodendrocytes have a feedback system that allows them to regulate myelin lipid synthesis depending on their myelinating state. The data presented are of potential clinical relevance as the human orthologs of the Zfp191 and MBP genes reside on a region of Chromosome 18 that is deleted in childhood leukodystrophies.

  8. Adrenocorticotropic hormone protects learning and memory function in epileptic Kcna1-null mice.

    PubMed

    Scantlebury, Morris H; Chun, Kyoung-Chul; Ma, Shun-Chieh; Rho, Jong M; Kim, Do Young

    2017-04-03

    ACTH, a member of the melanocortin family of peptides, is often used in the treatment of the developmental epileptic encephalopathy spectrum disorders including, Ohtahara, West, Lennox Gastaut and Landau-Kleffner Syndromes and electrical status epilepticus of sleep. In these disorders, although ACTH is often successful in controlling the seizures and/or inter-ictal EEG abnormalities, it is unknown whether ACTH possesses other beneficial effects independent of seizure control. We tested whether ACTH can ameliorate the intrinsic impairment of hippocampal-based learning and memory in epileptic Kcna1-null (KO) mice. We found that ACTH - administered in the form of Acthar Gel given i.p. four times daily at a dose of 4 IU/kg (16 IU/kg/day) for 7days - prevented impairment of long-term potentiation (LTP) evoked with high-frequency stimulation in CA1 hippocampus and also restored spatial learning and memory on the Barnes maze test. However, with this treatment regimen, ACTH did not exert a significant effect on the frequency of spontaneous recurrent seizures. Together, our findings indicate that ACTH can ameliorate memory impairment in epileptic Kcna1-null mice separate from seizure control, and suggest that this widely used peptide may exert direct nootropic effects in the epileptic brain.

  9. Transcriptional Fingerprint of Hypomyelination in Zfp191null and Shiverer (Mbpshi) Mice

    PubMed Central

    Aaker, Joshua D.; Elbaz, Benayahu; Wu, Yuwen; Looney, Timothy J.; Zhang, Li; Lahn, Bruce T.

    2016-01-01

    The transcriptional program that controls oligodendrocyte maturation and central nervous system (CNS) myelination has not been fully characterized. In this study, we use high-throughput RNA sequencing to analyze how the loss of a key transcription factor, zinc finger protein 191 (ZFP191), results in oligodendrocyte development abnormalities and CNS hypomyelination. Using a previously described mutant mouse that is deficient in ZFP191 protein expression (Zfp191null), we demonstrate that key transcripts are reduced in the whole brain as well as within oligodendrocyte lineage cells cultured in vitro. To determine whether the loss of myelin seen in Zfp191null mice contributes indirectly to these perturbations, we also examined the transcriptome of a well-characterized mouse model of hypomyelination, in which the myelin structural protein myelin basic protein (MBP) is deficient. Interestingly, Mbpshi (shiverer) mice had far fewer transcripts perturbed with the loss of myelin alone. This study demonstrates that the loss of ZFP191 disrupts expression of genes involved in oligodendrocyte maturation and myelination, largely independent from the loss of myelin. Nevertheless, hypomyelination in both mouse mutants results in the perturbation of lipid synthesis pathways, suggesting that oligodendrocytes have a feedback system that allows them to regulate myelin lipid synthesis depending on their myelinating state. The data presented are of potential clinical relevance as the human orthologs of the Zfp191 and MBP genes reside on a region of Chromosome 18 that is deleted in childhood leukodystrophies. PMID:27683878

  10. Dietary Carnosine Prevents Early Atherosclerotic Lesion Formation in ApoE-null Mice

    PubMed Central

    Barski, Oleg A.; Xie, Zhengzhi; Baba, Shahid P.; Sithu, Srinivas D.; Agarwal, Abhinav; Cai, Jian; Bhatnagar, Aruni; Srivastava, Sanjay

    2013-01-01

    Objective Atherosclerotic lesions are associated with the accumulation of reactive aldehydes derived from oxidized lipids. Although inhibition of aldehyde metabolism has been shown to exacerbate atherosclerosis and enhance the accumulation of aldehyde-modified proteins in atherosclerotic plaques, no therapeutic interventions have been devised to prevent aldehyde accumulation in atherosclerotic lesions. Approach and Results We examined the efficacy of carnosine, a naturally occurring β-alanyl-histidine dipeptide in preventing aldehyde toxicity and atherogenesis in apoE-null mice. In vitro, carnosine reacted rapidly with lipid peroxidation-derived unsaturated aldehydes. Gas chromatography mass-spectrometry analysis showed that carnosine inhibits the formation of free aldehydes - HNE and malonaldialdehyde in Cu2+-oxidized LDL. Preloading bone marrow-derived macrophages with cell-permeable carnosine analogs reduced HNE-induced apoptosis. Oral supplementation with octyl-D-carnosine decreased atherosclerotic lesion formation in aortic valves of apoE-null mice and attenuated the accumulation of protein-acrolein, protein-HHE and protein-HNE adducts in atherosclerotic lesions, while urinary excretion of aldehydes as carnosine conjugates was increased. Conclusions The results of this study suggest that carnosine inhibits atherogenesis by facilitating aldehyde removal from atherosclerotic lesions. Endogenous levels of carnosine may be important determinants of atherosclerotic lesion formation and treatment with carnosine or related peptides could be a useful therapy for the prevention or the treatment of atherosclerosis. PMID:23559625

  11. Perturbed desmosomal cadherin expression in grainy head-like 1-null mice

    PubMed Central

    Wilanowski, Tomasz; Caddy, Jacinta; Ting, Stephen B; Hislop, Nikki R; Cerruti, Loretta; Auden, Alana; Zhao, Lin-Lin; Asquith, Stephen; Ellis, Sarah; Sinclair, Rodney; Cunningham, John M; Jane, Stephen M

    2008-01-01

    In Drosophila, the grainy head (grh) gene plays a range of key developmental roles through the regulation of members of the cadherin gene family. We now report that mice lacking the grh homologue grainy head-like 1 (Grhl1) exhibit hair and skin phenotypes consistent with a reduction in expression of the genes encoding the desmosomal cadherin, desmoglein 1 (Dsg1). Grhl1-null mice show an initial delay in coat growth, and older mice exhibit hair loss as a result of poor anchoring of the hair shaft in the follicle. The mice also develop palmoplantar keratoderma, analogous to humans with DSG1 mutations. Sequence analysis, DNA binding, and chromatin immunoprecipitation experiments demonstrate that the human and mouse Dsg1 promoters are direct targets of GRHL1. Ultrastructural analysis reveals reduced numbers of abnormal desmosomes in the interfollicular epidermis. These findings establish GRHL1 as an important regulator of the Dsg1 genes in the context of hair anchorage and epidermal differentiation, and suggest that cadherin family genes are key targets of the grainy head-like genes across 700 million years of evolution. PMID:18288204

  12. Nitric oxide synthase mediates the ability of darbepoetin alpha to improve the cognitive performance of STOP null mice.

    PubMed

    Kajitani, Kosuke; Thorne, Michael; Samson, Michel; Robertson, George S

    2010-07-01

    STOP (stable tubule only polypeptide) null mice display neurochemical and behavioral abnormalities that resemble several well-recognized features of schizophrenia. Recent evidence suggests that the hematopoietic growth factor erythropoietin improves the cognitive performance of schizophrenics. The mechanism, however, by which erythropoietin is able to improve the cognition of schizophrenics is unclear. To address this question, we first determined whether acute administration of the erythropoietin analog known as darbepoetin alpha (D. alpha) improved performance deficits of STOP null mice in the novel objective recognition task (NORT). NORT performance of STOP null mice, but not wild-type littermates, was enhanced 3 h after a single injection of D. alpha (25 microg/kg, i.p.). Improved NORT performance was accompanied by elevated NADPH diaphorase staining in the ventral hippocampus as well as medial and cortical aspects of the amygdala, indicative of increased nitric oxide synthase (NOS) activity in these structures. NOS generates the intracellular messenger nitric oxide (NO) implicated in learning and memory. In keeping with this hypothesis, D. alpha significantly increased NO metabolite levels (nitrate and nitrite, NOx) in the hippocampus of both wild-type and STOP null mice. The NOS inhibitor, N (G)-nitro-L- arginine methyl ester (L-NAME; 25 mg/kg, i.p.), completely reversed the increase in hippocampal NOx levels produced by D. alpha. Moreover, L-NAME also inhibited the ability of D. alpha to improve the NORT performance of STOP null mice. Taken together, these observations suggest D. alpha enhances the NORT performance of STOP null mice by increasing production of NO.

  13. Vitamin D and Human Health: Lessons from Vitamin D Receptor Null Mice

    PubMed Central

    Bouillon, Roger; Carmeliet, Geert; Verlinden, Lieve; van Etten, Evelyne; Verstuyf, Annemieke; Luderer, Hilary F.; Lieben, Liesbet; Mathieu, Chantal; Demay, Marie

    2008-01-01

    The vitamin D endocrine system is essential for calcium and bone homeostasis. The precise mode of action and the full spectrum of activities of the vitamin D hormone, 1,25-dihydroxyvitamin D [1,25-(OH)2D], can now be better evaluated by critical analysis of mice with engineered deletion of the vitamin D receptor (VDR). Absence of a functional VDR or the key activating enzyme, 25-OHD-1α-hydroxylase (CYP27B1), in mice creates a bone and growth plate phenotype that mimics humans with the same congenital disease or severe vitamin D deficiency. The intestine is the key target for the VDR because high calcium intake, or selective VDR rescue in the intestine, restores a normal bone and growth plate phenotype. The VDR is nearly ubiquitously expressed, and almost all cells respond to 1,25-(OH)2D exposure; about 3% of the mouse or human genome is regulated, directly and/or indirectly, by the vitamin D endocrine system, suggesting a more widespread function. VDR-deficient mice, but not vitamin D- or 1α-hydroxylase-deficient mice, and man develop total alopecia, indicating that the function of the VDR and its ligand is not fully overlapping. The immune system of VDR- or vitamin D-deficient mice is grossly normal but shows increased sensitivity to autoimmune diseases such as inflammatory bowel disease or type 1 diabetes after exposure to predisposing factors. VDR-deficient mice do not have a spontaneous increase in cancer but are more prone to oncogene- or chemocarcinogen-induced tumors. They also develop high renin hypertension, cardiac hypertrophy, and increased thrombogenicity. Vitamin D deficiency in humans is associated with increased prevalence of diseases, as predicted by the VDR null phenotype. Prospective vitamin D supplementation studies with multiple noncalcemic endpoints are needed to define the benefits of an optimal vitamin D status. PMID:18694980

  14. Reduced wheel running and blunted effects of voluntary exercise in LPA1-null mice: The importance of assessing the amount of running in transgenic mice studies

    PubMed Central

    Castilla-Ortega, Estela; Rosell-Valle, Cristina; Blanco, Eduardo; Pedraza, Carmen; Chun, Jerold; de Fonseca, Fernando Rodríguez; Estivill-Torrús, Guillermo; Santín, Luis J.

    2014-01-01

    This work was aimed to assess whether voluntary exercise rescued behavioral and hippocampal alterations in mice lacking the lysophosphatidic acid LPA1 receptor (LPA1-null mice), studying the potential relationship between the amount of exercise performed and its effects. Normal and LPA1-null mice underwent 23 days of free wheel running and were tested for open-field behavior and adult hippocampal neurogenesis (cell proliferation, immature neurons, cell survival). Running decreased anxiety-like behavior in both genotypes but increased exploration only in the normal mice. While running affected all neurogenesis-related measures in normal mice (especially in the suprapyramidal blade of the dentate gyrus), only a moderate increase in cell survival was found in the mutants. Importantly, the LPA1-nulls showed notably reduced running. Analysis suggested that defective running in the LPA1-null mice could contribute to explain the scarce benefit of the voluntary exercise treatment. On the other hand, a literature review revealed that voluntary exercise is frequently used to modulate behavior and the hippocampus in transgenic mice, but half of the studies did not assess the quantity of running, overlooking any potential running impairments. This study adds evidence to the relevance of the quantity of exercise performed, emphasizing the importance of its assessment in transgenic mice research. PMID:24055600

  15. A human apolipoprotein E mimetic peptide reduces atherosclerosis in aged apolipoprotein E null mice

    PubMed Central

    Xu, Yanyong; Liu, Hongmei; Liu, Mengting; Li, Feifei; Liu, Liangchen; Du, Fen; Fan, Daping; Yu, Hong

    2016-01-01

    Apolipoprotein E (apoE) is well known as an antiatherogenic protein via regulating lipid metabolism and inflammation. We previously reported that a human apoE mimetic peptide, EpK, reduced atherosclerosis in apoE null (apoE-/-) mice through reducing inflammation without affecting plasma lipid levels. Here, we construct another human apoE mimetic peptide, named hEp, and investigate whether expression of hEp can reduce atherosclerotic lesion development in aged female apoE-/- mice with pre-existing lesions. We found that chemically synthesized hEp significantly decreased cholesterol accumulation induced by oxidized low density lipoprotein and the expression of inflammatory cytokines TNFα and IL-6 induced by lipopolysaccharide in macrophages. In an in vivo study, Lv-hEp-GFP lentiviruses were intravenously injected into 9 month-old apoE-/- mice. Mice were then fed a chow diet for 18 weeks. Results showed that in comparison to the Lv-GFP lentivirus injection (Lv-GFP) group, Lv-hEp-GFP lentivirus injection achieved hepatic hEp expression and secretion in apoE-/- mice. It was observed that hEp expression significantly reduced plasma VLDL and LDL cholesterol levels and decreased aortic atherosclerotic lesions. This was accompanied by an increase of LDL receptor expression and a reduction of TNFα and IL-6 mRNA levels in the liver. Moreover, expression of hEp increased plasma paraoxonase-1 activity and decreased plasma myeloperoxidase activity and serum amyloid A levels. Our study provides evidence that hEp may be developed as a promising therapeutic apoE mimetic peptide for atherosclerosis-related cardiovascular diseases through its induction of plasma VLDL/LDL cholesterol clearance as well as its anti-oxidative and anti-inflammatory activities. PMID:27648138

  16. Gamma Interferon (IFN-γ) Receptor Null-Mutant Mice Are More Susceptible to Herpes Simplex Virus Type 1 Infection than IFN-γ Ligand Null-Mutant Mice

    PubMed Central

    Cantin, Edouard; Tanamachi, Becky; Openshaw, Harry; Mann, Jeff; Clarke, Ken

    1999-01-01

    Mouse strains with null mutations in the gamma interferon gene (Ifng) or the gamma interferon receptor gene (Ifngr) have been engineered. The use of these strains as animal models of viral and bacterial infections has enhanced our understanding of the role of gamma interferon (IFN-γ) in the host immune response. However, direct comparisons between Ifng−/− (GKO) and Ifngr−/− (RGKO) mice have been problematic because previously available strains of these mice have had different genetic backgrounds (i.e., C57BL/6 and BALB/c for GKO mice and 129/Sv//Ev for RGKO mice). To enable direct comparison of herpes simplex virus type 1 (HSV-1) infections in GKO and RGKO mice, we introduced the IFN-γ null mutation into the 129/Sv//Ev background. We report that, after HSV-1 inoculation, mortality was significantly greater in RGKO mice than in GKO mice (38 versus 23%, P = 0.0001). Similarly, the mortality from vaccinia virus challenge was significantly greater in RGKO mice than in GKO mice. With differences in genetic background excluded as a confounding issue, these results are consistent with the existence of an alternative ligand(s) for the IFN-γ receptor that is also capable of mediating protection against viral challenge. PMID:10233988

  17. Hyaluronan Synthase 3 Null Mice Exhibit Decreased Intestinal Inflammation and Tissue Damage in the DSS-Induced Colitis Model

    PubMed Central

    Kessler, Sean P.; Obery, Dana R.; de la Motte, Carol

    2015-01-01

    Hyaluronan (HA) overproduction is a hallmark of multiple inflammatory diseases, including inflammatory bowel disease (IBD). Hyaluronan can act as a leukocyte recruitment molecule and in the most common mouse model of intestinal inflammation, the chemically induced dextran sodium sulfate (DSS) experimental colitis model, we previously determined that changes in colon distribution of HA occur before inflammation. Therefore, we hypothesized that, during a pathologic challenge, HA promotes inflammation. In this study, we tested the progression of inflammation in mice null for the hyaluronan synthase genes (HAS1, HAS3, or both HAS1 and HAS3) in the DSS-colitis model. Our data demonstrate that both the HAS1/HAS3 double and the HAS3 null mice are protected from colitis, compared to wild-type and HAS1 null mice, as determined by measurement of weight loss, disease activity, serum IL-6 levels, histologic scoring, and immunohistochemistry. Most notable is the dramatic increase in submucosal microvasculature, hyaluronan deposition, and leukocyte infiltration in the inflamed colon tissue of wild-type and HAS1 null mice. Our data suggest, HAS3 plays a crucial role in driving gut inflammation. Developing a temporary targeted therapeutic intervention of HAS3 expression or function in the microcirculation may emerge as a desirable strategy toward tempering colitis in patients undergoing flares of IBD. PMID:26448758

  18. Altered Morphine-Induced Analgesia in Neurotensin Type 1 Receptor Null Mice

    PubMed Central

    Roussy, Geneviève; Beaudry, Hélène; Lafrance, Mylène; Belleville, Karine; Beaudet, Nicolas; Wada, Keiji; Gendron, Louis; Sarret, Philippe

    2013-01-01

    Both neurotensin (NT) and opioid agonists have been shown to induce antinociception in rodents after central administration. Besides, previous studies have revealed the existence of functional interactions between NT and opioid systems in the regulation of pain processing. We recently demonstrated that NTS1 receptors play a key role in the mediation of the analgesic effects of NT in long-lasting pain. In the present study, we therefore investigated whether NTS1 gene deletion affected the antinociceptive action of mu opioid drugs. To this end, pain behavioral responses to formalin were determined following systemic administration of morphine in both male and female NTS1 knockout mice. Acute injection of morphine (2 or 5 mg/kg) produced strong antinociceptive effects in both male and female wild-type littermates, with no significant sex differences. On the other hand, morphine analgesia was considerably reduced in NTS1-deficient mice of both sexes compared to their respective controls, indicating that the NTS1 receptor actively participates in mu opioid alleviating pain. By examining specifically the flinching, licking and biting nociceptive behaviors, we also showed that the functional crosstalk between NTS1 and mu opioid receptors influences the supraspinally-mediated behaviors. Interestingly, sexual dimorphic action of morphine-induced pain inhibition was found in NTS1 null mice in the formalin test, suggesting that the endogenous NT system interacts differently with the opioid network in male and female mice. Altogether, these results demonstrated that NTS1 receptor activation operates downstream to the opioidergic transmission and that NTS1-selective agonists combined with morphine may act synergistically to reduce persistent pain. PMID:20727387

  19. Calcium Homeostasis and Muscle Energy Metabolism Are Modified in HspB1-Null Mice

    PubMed Central

    Picard, Brigitte; Kammoun, Malek; Gagaoua, Mohammed; Barboiron, Christiane; Meunier, Bruno; Chambon, Christophe; Cassar-Malek, Isabelle

    2016-01-01

    Hsp27—encoded by HspB1—is a member of the small heat shock proteins (sHsp, 12–43 kDa (kilodalton)) family. This protein is constitutively present in a wide variety of tissues and in many cell lines. The abundance of Hsp27 is highest in skeletal muscle, indicating a crucial role for muscle physiology. The protein identified as a beef tenderness biomarker was found at a crucial hub in a functional network involved in beef tenderness. The aim of this study was to analyze the proteins impacted by the targeted invalidation of HspB1 in the Tibialis anterior muscle of the mouse. Comparative proteomics using two-dimensional gel electrophoresis revealed 22 spots that were differentially abundant between HspB1-null mice and their controls that could be identified by mass spectrometry. Eighteen spots were more abundant in the muscle of the mutant mice, and four were less abundant. The proteins impacted by the absence of Hsp27 belonged mainly to calcium homeostasis (Srl and Calsq1), contraction (TnnT3), energy metabolism (Tpi1, Mdh1, PdhB, Ckm, Pygm, ApoA1) and the Hsp proteins family (HspA9). These data suggest a crucial role for these proteins in meat tenderization. The information gained by this study could also be helpful to predict the side effects of Hsp27 depletion in muscle development and pathologies linked to small Hsps. PMID:28248227

  20. Deficiency in acellular cementum and periodontal attachment in bsp null mice.

    PubMed

    Foster, B L; Soenjaya, Y; Nociti, F H; Holm, E; Zerfas, P M; Wimer, H F; Holdsworth, D W; Aubin, J E; Hunter, G K; Goldberg, H A; Somerman, M J

    2013-02-01

    Bone sialoprotein (BSP) is an extracellular matrix protein found in mineralized tissues of the skeleton and dentition. BSP is multifunctional, affecting cell attachment and signaling through an RGD integrin-binding region, and acting as a positive regulator for mineral precipitation by nucleating hydroxyapatite crystals. BSP is present in cementum, the hard tissue covering the tooth root that anchors periodontal ligament (PDL) attachment. To test our hypothesis that BSP plays an important role in cementogenesis, we analyzed tooth development in a Bsp null ((-/-)) mouse model. Developmental analysis by histology, histochemistry, and SEM revealed a significant reduction in acellular cementum formation on Bsp (-/-) mouse molar and incisor roots, and the cementum deposited appeared hypomineralized. Structural defects in cementum-PDL interfaces in Bsp (-/-) mice caused PDL detachment, likely contributing to the high incidence of incisor malocclusion. Loss of BSP caused progressively disorganized PDL and significantly increased epithelial down-growth with aging. Bsp (-/-) mice displayed extensive root and alveolar bone resorption, mediated by increased RANKL and the presence of osteoclasts. Results collected here suggest that BSP plays a non-redundant role in acellular cementum formation, likely involved in initiating mineralization on the root surface. Through its importance to cementum integrity, BSP is essential for periodontal function.

  1. Deficiency in Acellular Cementum and Periodontal Attachment in Bsp Null Mice

    PubMed Central

    Foster, B.L.; Soenjaya, Y.; Nociti, F.H.; Holm, E.; Zerfas, P.M.; Wimer, H.F.; Holdsworth, D.W.; Aubin, J.E.; Hunter, G.K.; Goldberg, H.A.; Somerman, M.J.

    2012-01-01

    Bone sialoprotein (BSP) is an extracellular matrix protein found in mineralized tissues of the skeleton and dentition. BSP is multifunctional, affecting cell attachment and signaling through an RGD integrin-binding region, and acting as a positive regulator for mineral precipitation by nucleating hydroxyapatite crystals. BSP is present in cementum, the hard tissue covering the tooth root that anchors periodontal ligament (PDL) attachment. To test our hypothesis that BSP plays an important role in cementogenesis, we analyzed tooth development in a Bsp null (-/-) mouse model. Developmental analysis by histology, histochemistry, and SEM revealed a significant reduction in acellular cementum formation on Bsp-/- mouse molar and incisor roots, and the cementum deposited appeared hypomineralized. Structural defects in cementum-PDL interfaces in Bsp-/- mice caused PDL detachment, likely contributing to the high incidence of incisor malocclusion. Loss of BSP caused progressively disorganized PDL and significantly increased epithelial down-growth with aging. Bsp-/- mice displayed extensive root and alveolar bone resorption, mediated by increased RANKL and the presence of osteoclasts. Results collected here suggest that BSP plays a non-redundant role in acellular cementum formation, likely involved in initiating mineralization on the root surface. Through its importance to cementum integrity, BSP is essential for periodontal function. PMID:23183644

  2. Cognition and Mood-Related Behaviors in L3mbtl1 Null Mutant Mice

    PubMed Central

    Shen, Erica Y.; Jiang, Yan; Mao, Wenjie; Futai, Kensuke; Hock, Hanno; Akbarian, Schahram

    2015-01-01

    Alterations in histone lysine methylation and epigenetic regulators of gene expression could play a role in the neurobiology and treatment of patients diagnosed with mood spectrum disorder, including depression and anxiety. Mutations and altered expression of various lysine methyltransferases (KMTs) and demethylases (KDMs) have been linked to changes in motivational and emotional behaviors in preclinical model systems. However, it is not known whether regulators operating downstream of histone lysine methylation could affect mood-related behavior. Malignant Brain Tumor (MBT) domain ‘chromatin reader’ proteins bind to methylated histone lysine residues and associate with chromatin remodeling complexes to facilitate or repress gene expression. MBT proteins, including the founding member, L3mbtl1, maintain high levels of expression in neurons of the mature brain. Here, we exposed L3mbtl1 null mutant mice to a wide range of tests exploring cognition and mood-relevant behaviors at baseline and in the context of social isolation, as a stressor to elicit depression-related behavior in susceptible mice. L3mbtl1 loss-of-function was associated with significant decreases in depression and and anxiety in some of the behavioral paradigms. This was not associated with a more generalized neurological dysfunction because cognition and memory remained unaltered in comparison to controls. These findings warrant further investigations on the role of MBT chromatin reader proteins in the context of emotional and affective behaviors. PMID:25849281

  3. Taurodontism, variations in tooth number, and misshapened crowns in Wnt10a null mice and human kindreds

    PubMed Central

    Yang, Jie; Wang, Shih-Kai; Choi, Murim; Reid, Bryan M; Hu, Yuanyuan; Lee, Yuan-Ling; Herzog, Curtis R; Kim-Berman, Hera; Lee, Moses; Benke, Paul J; Kent Lloyd, K C; Simmer, James P; Hu, Jan C-C

    2015-01-01

    WNT10A is a signaling molecule involved in tooth development, and WNT10A defects are associated with tooth agenesis. We characterized Wnt10a null mice generated by the knockout mouse project (KOMP) and six families with WNT10A mutations, including a novel p.Arg104Cys defect, in the absence of EDA,EDAR, or EDARADD variations. Wnt10a null mice exhibited supernumerary mandibular fourth molars, and smaller molars with abnormal cusp patterning and root taurodontism. Wnt10a−/− incisors showed distinctive apical–lingual wedge-shaped defects. These findings spurred us to closely examine the dental phenotypes of our WNT10A families. WNT10A heterozygotes exhibited molar root taurodontism and mild tooth agenesis (with incomplete penetrance) in their permanent dentitions. Individuals with two defective WNT10A alleles showed severe tooth agenesis and had fewer cusps on their molars. The misshapened molar crowns and roots were consistent with the Wnt10a null phenotype and were not previously associated with WNT10A defects. The missing teeth contrasted with the presence of supplemental teeth in the Wnt10a null mice and demonstrated mammalian species differences in the roles of Wnt signaling in early tooth development. We conclude that molar crown and root dysmorphologies are caused by WNT10A defects and that the severity of the tooth agenesis correlates with the number of defective WNT10A alleles. PMID:25629078

  4. Hexim1 heterozygosity stabilizes atherosclerotic plaque and decreased steatosis in ApoE null mice fed atherogenic diet.

    PubMed

    Dhar-Mascareno, Manya; Rozenberg, Inna; Iqbal, Jahangir; Hussain, M Mahmood; Beckles, Daniel; Mascareno, Eduardo

    2017-02-01

    Hexim-1 is an inhibitor of RNA polymerase II transcription elongation. Decreased Hexim-1 expression in animal models of chronic diseases such as left ventricular hypertrophy, obesity and cancer triggered significant changes in adaptation and remodeling. The main aim of this study was to evaluate the role of Hexim1 in lipid metabolism focused in the progression of atherosclerosis and steatosis. We used the C57BL6 apolipoprotein E (ApoE null) crossed bred to C57BL6Hexim1 heterozygous mice to obtain ApoE null - Hexim1 heterozygous mice (ApoE-HT). Both ApoE null backgrounds were fed high fat diet for twelve weeks. Then, we evaluated lipid metabolism, atherosclerotic plaque formation and liver steatosis. In order to understand changes in the transcriptome of both backgrounds during the progression of steatosis, we performed Affymetrix mouse 430 2.0 microarray. After 12 weeks of HFD, ApoE null and ApoE-HT showed similar increase of cholesterol and triglycerides in plasma. Plaque composition was altered in ApoE-HT. Additionally, liver triglycerides and steatosis were decreased in ApoE-HT mice. Affymetrix analysis revealed that decreased steatosis might be due to impaired inducible SOCS3 expression in ApoE-HT mice. In conclusion, decreased Hexim-1 expression does not alter cholesterol metabolism in ApoE null background after HFD. However, it promotes stable atherosclerotic plaque and decreased steatosis by promoting the anti-inflammatory TGFβ pathway and blocking the expression of the inducible and pro-inflammatory expression of SOCS3 respectively.

  5. Toll-Like Receptor 4 Mutant and Null Mice Retain Morphine-Induced Tolerance, Hyperalgesia, and Physical Dependence

    PubMed Central

    Mattioli, Theresa Alexandra; Leduc-Pessah, Heather; Skelhorne-Gross, Graham; Nicol, Christopher J. B.; Milne, Brian; Trang, Tuan; Cahill, Catherine M.

    2014-01-01

    The innate immune system modulates opioid-induced effects within the central nervous system and one target that has received considerable attention is the toll-like receptor 4 (TLR4). Here, we examined the contribution of TLR4 in the development of morphine tolerance, hyperalgesia, and physical dependence in two inbred mouse strains: C3H/HeJ mice which have a dominant negative point mutation in the Tlr4 gene rendering the receptor non-functional, and B10ScNJ mice which are TLR4 null mutants. We found that neither acute antinociceptive response to a single dose of morphine, nor the development of analgesic tolerance to repeated morphine treatment, was affected by TLR4 genotype. Likewise, opioid induced hyperalgesia and opioid physical dependence (assessed by naloxone precipitated withdrawal) were not altered in TLR4 mutant or null mice. We also examined the behavioural consequence of two stereoisomers of naloxone: (−) naloxone, an opioid receptor antagonist, and (+) naloxone, a purported antagonist of TLR4. Both stereoisomers of naloxone suppressed opioid induced hyperalgesia in wild-type control, TLR4 mutant, and TLR4 null mice. Collectively, our data suggest that TLR4 is not required for opioid-induced analgesic tolerance, hyperalgesia, or physical dependence. PMID:24824631

  6. Behavioral phenotype of maLPA1-null mice: increased anxiety-like behavior and spatial memory deficits

    PubMed Central

    Santin, L.J.; Bilbao, A.; Pedraza, C.; Matas-Rico, E.; López-Barroso, D.; Castilla-Ortega, E.; Sánchez-López, J.; Riquelme, R.; Varela-Nieto, I.; de la Villa, P.; Suardíaz, M.; Chun, J.; De Fonseca, F. Rodriguez; Estivill-Torrús, G.

    2016-01-01

    Lysophosphatidic acid (LPA) has emerged as a new regulatory molecule in the brain. Recently, some studies have demonstrated a role for this molecule and its LPA1 receptor in the regulation of plasticity and neurogenesis in the adult brain. However, no systematic studies have been conducted to investigate whether the LPA1 receptor is involved in behavior. Here we studied the phenotype of maLPA1–null mice, which bear a targeted deletion at the lpa1 locus, in a battery of tests examining neurologic performance, habituation in exploratory behavior in response to low and mild anxiety environments and spatial memory. MaLPA1-null mutants showed deficits in both olfaction and somesthesis, but not in retinal or auditory functions. Sensorimotor coordination was impaired only in the equilibrium and grasping reflexes. The mice also showed impairments in neuromuscular strength and analgesic response. No additional differences were observed in the rest of the tests used to study sensoriomotor orientation, limb reflexes, and coordinated limb use. At behavioral level, maLPA1-null mice showed an impaired exploration in the open field and increased anxiety-like response when exposed to the elevated plus maze. Furthermore, the mice exhibit impaired spatial memory retention and reduced use of spatial strategies in the Morris water maze. We propose that the LPA1 receptor may play a major role in both spatial memory and response to anxiety-like conditions. PMID:19689455

  7. Nephropathy in Pparg-null mice highlights PPARγ systemic activities in metabolism and in the immune system

    PubMed Central

    Toffoli, Barbara; Gilardi, Federica; Winkler, Carine; Soderberg, Magnus; Kowalczuk, Laura; Arsenijevic, Yvan; Bamberg, Krister; Bonny, Olivier

    2017-01-01

    Peroxisome proliferator-activated receptor γ (PPARγ) is a ligand-dependent transcription factor involved in many aspects of metabolism, immune response, and development. Total-body deletion of the two Pparg alleles provoked generalized lipoatrophy along with severe type 2 diabetes. Herein, we explore the appearance and development of structural and functional alterations of the kidney, comparing Pparg null-mice to their littermate controls (carrying Pparg floxed alleles). We show that renal hypertrophy and functional alterations with increased glucosuria and albuminuria are already present in 3 weeks-old Pparg null-mice. Renal insufficiency with decreased creatinine clearance progress at 7 weeks of age, with the advance of the type 2 diabetes. At 52 weeks of age, these alterations are accompanied by signs of fibrosis and mesangial expansion. More intriguingly, aged Pparg null-mice concomitantly present an anti-phospholipid syndrome (APS), characterized by the late appearance of microthrombi and a mesangioproliferative pattern of glomerular injury, associated with significant plasmatic levels of anti-β2- glycoprotein1 antibodies and renal deposition of IgG, IgM, and C3. Thus, in line with the role of PPARγ in metabolic homeostasis, Pparg null-mice first represent a potent model for studying the initiation and the development of diabetic nephropathy. Second, and in relation with the important PPARγ activity in inflammation and in immune system, these mice also highlight a new role for PPARγ signaling in the promotion of APS, a syndrome whose pathogenesis is poorly known and whose current treatment is limited to prevention of thrombosis events. PMID:28182703

  8. Antenatal maternal low protein diet: ACE-2 in the mouse lung and sexually dimorphic programming of hypertension.

    PubMed

    Goyal, Ravi; Van-Wickle, Jonathan; Goyal, Dipali; Longo, Lawrence D

    2015-05-14

    Elevated blood pressure is an important global health problem, and in-utero under-nutrition may be an important factor in the pathogenesis of hypertension. In the present study, we tested the hypothesis that antenatal maternal low protein diet (MLPD) leads to sexually dimorphic developmental programming of the components of the pulmonary renin-angiotensin system. This may be important in the antenatal MLPD-associated development of hypertension. In pregnant mice, we administered normal (control) and isocaloric 50% protein restricted diet, commencing one week before mating and continuing until delivery of the pups. From the 18th to 24th week postnatal, we measured blood pressure in the offspring by use of a non-invasive tail-cuff method. In the same mice, we examined the mRNA and protein expression of the key components of the pulmonary renin-angiotensin system. Also, we examined microRNA complementary to angiotensin converting enzymes (ACE) 2 in the offspring lungs. Our results demonstrate that as a consequence of antenatal MLPD: 1) pup birthweight was significantly reduced in both sexes. 2) female offspring developed hypertension, but males did not. 3) In female offspring, ACE-2 protein expression was significantly reduced without any change in the mRNA levels. 4) miRNA 429, which has a binding site on ACE-2 - 3' UTR was significantly upregulated in the female antenatal MLPD offspring. 5) In males, ACE-2 mRNA and protein expression were unaltered. We conclude that in the mouse, antenatal MLPD-induced reduction of ACE-2 in the female offspring lung may be an important mechanisms in sexually dimorphic programming of hypertension.

  9. Increased maternofetal calcium flux in parathyroid hormone-related protein-null mice

    PubMed Central

    Bond, H; Dilworth, M R; Baker, B; Cowley, E; Requena Jimenez, A; Boyd, R D H; Husain, S M; Ward, B S; Sibley, C P; Glazier, J D

    2008-01-01

    The role of parathyroid hormone-related protein (PTHrP) in fetal calcium homeostasis and placental calcium transport was examined in mice homozygous for the deletion of the PTHrP gene (PTHrP−/− null; NL) compared to PTHrP+/+ (wild-type; WT) and PTHrP+/− (heterozygous; HZ) littermates. Fetal blood ionized calcium was significantly reduced in NL fetuses compared to WT and HZ groups at 18 days of pregnancy (dp) with abolition of the fetomaternal calcium gradient. In situ placental perfusion of the umbilical circulation at 18 dp was used to measure unidirectional clearance of 45Ca across the placenta in maternofetal (CaKmf) and fetoplacental (CaKfp) directions; CaKfp was < 5% of CaKmf for all genotypes. At 18 dp, CaKmf across perfused placenta and intact placenta (CaKmf(intact)) were similar and concordant with net calcium accretion rates in vivo. CaKmf was significantly raised in NL fetuses compared to WT and HZ littermates. Calcium accretion was significantly elevated in NL fetuses by 19 dp. Placental calbindin-D9K expression in NL fetuses was marginally enhanced (P < 0.07) but expression of TRPV6/ECaC2 and plasma membrane Ca2+-ATPase (PMCA) isoforms 1 and 4 were unaltered. We conclude that PTHrP is an important regulator of fetal calcium homeostasis with its predominant effect being on unidirectional maternofetal transfer, probably mediated by modifying placental calbindin-D9K expression. In situ perfusion of mouse placenta is a robust methodology for allowing detailed dissection of placental transfer mechanisms in genetically modified mice. PMID:18258656

  10. Mecp2-Null Mice Provide New Neuronal Targets for Rett Syndrome

    PubMed Central

    Urdinguio, Rocio G.; Lopez-Serra, Lidia; Lopez-Nieva, Pilar; Alaminos, Miguel; Diaz-Uriarte, Ramon; Fernandez, Agustin F.; Esteller, Manel

    2008-01-01

    Background Rett syndrome (RTT) is a complex neurological disorder that is one of the most frequent causes of mental retardation in women. A great landmark in research in this field was the discovery of a relationship between the disease and the presence of mutations in the gene that codes for the methyl-CpG binding protein 2 (MeCP2). Currently, MeCP2 is thought to act as a transcriptional repressor that couples DNA methylation and transcriptional silencing. The present study aimed to identify new target genes regulated by Mecp2 in a mouse model of RTT. Methodology/Principal Findings We have compared the gene expression profiles of wild type (WT) and Mecp2-null (KO) mice in three regions of the brain (cortex, midbrain, and cerebellum) by using cDNA microarrays. The results obtained were confirmed by quantitative real-time PCR. Subsequent chromatin immunoprecipitation assays revealed seven direct target genes of Mecp2 bound in vivo (Fkbp5, Mobp, Plagl1, Ddc, Mllt2h, Eya2, and S100a9), and three overexpressed genes due to an indirect effect of a lack of Mecp2 (Irak1, Prodh and Dlk1). The regions bound by Mecp2 were always methylated, suggesting the involvement of the methyl-CpG binding domain of the protein in the mechanism of interaction. Conclusions We identified new genes that are overexpressed in Mecp2-KO mice and are excellent candidate genes for involvement in various features of the neurological disease. Our results demonstrate new targets of MeCP2 and provide us with a better understanding of the underlying mechanisms of RTT. PMID:18989361

  11. Constitutive Androgen Receptor-Null Mice Are Sensitive to the Toxic Effects of Parathion: Association with Reduced Cytochrome P450-Mediated Parathion MetabolismS⃞

    PubMed Central

    Mota, Linda C.; Hernandez, Juan P.

    2010-01-01

    Constitutive androgen receptor (CAR) is activated by several chemicals and in turn regulates multiple detoxification genes. Our research demonstrates that parathion is one of the most potent, environmentally relevant CAR activators with an EC50 of 1.43 μM. Therefore, animal studies were conducted to determine whether CAR was activated by parathion in vivo. Surprisingly, CAR-null mice, but not wild-type (WT) mice, showed significant parathion-induced toxicity. However, parathion did not induce Cyp2b expression, suggesting that parathion is not a CAR activator in vivo, presumably because of its short half-life. CAR expression is also associated with the expression of several drug-metabolizing cytochromes P450 (P450). CAR-null mice demonstrate lower expression of Cyp2b9, Cyp2b10, Cyp2c29, and Cyp3a11 primarily, but not exclusively in males. Therefore, we incubated microsomes from untreated WT and CAR-null mice with parathion in the presence of esterase inhibitors to determine whether CAR-null mice show perturbed P450-mediated parathion metabolism compared with that in WT mice. The metabolism of parathion to paraoxon and p-nitrophenol (PNP) was reduced in CAR-null mice with male CAR-null mice showing reduced production of both paraoxon and PNP, and female CAR-null mice showing reduced production of only PNP. Overall, the data indicate that CAR-null mice metabolize parathion slower than WT mice. These results provide a potential mechanism for increased sensitivity of individuals with lower CAR activity such as newborns to parathion and potentially other chemicals due to decreased metabolic capacity. PMID:20573718

  12. ACE2/Ang 1-7 axis: A critical regulator of epicardial adipose tissue inflammation and cardiac dysfunction in obesity

    PubMed Central

    Patel, Vaibhav B.; Basu, Ratnadeep; Oudit, Gavin Y.

    2016-01-01

    ABSTRACT Obesity is characterized by an excessive fat accumulation in adipose tissues leading to weight gain and is increasing in prevalence and is strongly associated with metabolic and cardiovascular disorders. The renin-angiotensin system (RAS) has emerged as a key pathogenic mechanism for these disorders; activated RAS and angiotensin (Ang) II production results in worsening of cardiovascular diseases and angiotensin converting enzyme 2 (ACE2) negatively regulates RAS by metabolizing Ang II into Ang 1-7. ACE2 is expressed in the adipocytes and its expression is upregulated in response to high fat diet induced obesity in mice. Loss of ACE2 results in heart failure with preserved ejection fraction which is mediated in part by epicardial adipose tissue inflammation. Angiotensin 1-7 reduces the obesity associated cardiac dysfunction predominantly via its role in adiponectin expression and attenuation of epicardial adipose tissue inflammation. Human heart disease is also linked with inflammed epicardial adipose tissue. Here, we discuss the important interpretation of the novel of ACE2/Ang 1-7 pathway in obesity associated cardiac dysfunction. PMID:27617176

  13. Lumbar intervertebral disc degeneration associated with axial and radiating low back pain in ageing SPARC-null mice.

    PubMed

    Millecamps, Magali; Tajerian, Maral; Naso, Lina; Sage, E Helene; Stone, Laura S

    2012-06-01

    Chronic low back pain (LBP) is a complex, multifactorial disorder with unclear underlying mechanisms. In humans and rodents, decreased expression of secreted protein acidic rich in cysteine (SPARC) is associated with intervertebral disc (IVD) degeneration and signs of LBP. The current study investigates the hypothesis that IVD degeneration is a risk factor for chronic LBP. SPARC-null and age-matched control mice ranging from 6 to 78 weeks of age were evaluated in this study. X-ray and histologic analysis revealed reduced IVD height, increased wedging, and signs of degeneration (bulging and herniation). Cutaneous sensitivity to cold, heat, and mechanical stimuli were used as measures of referred (low back and tail) and radiating pain (hind paw). Region specificity was assessed by measuring icilin- and capsaicin-evoked behaviour after subcutaneous injection into the hind paw or upper lip. Axial discomfort was measured by the tail suspension and grip force assays. Motor impairment was determined by the accelerating rotarod. Physical function was evaluated by voluntary activity after axial strain or during ambulation with forced lateral flexion. SPARC-null mice developed (1) region-specific, age-dependent hypersensitivity to cold, icilin, and capsaicin (hind paw only), (2) axial discomfort, (3) motor impairment, and (4) reduced physical function. Morphine (6 mg/kg, i.p.) reduced cutaneous sensitivity and alleviated axial discomfort in SPARC-null mice. Ageing SPARC-null mice mirror many aspects of the complex and challenging nature of LBP in humans and incorporate both anatomic and functional components of the disease. The current study supports the hypothesis that IVD degeneration is a risk factor for chronic LBP.

  14. A mouse renin distal enhancer is essential for blood pressure homeostasis in BAC-rescued renin-null mutant mice.

    PubMed

    Tanimoto, Keiji; Kanafusa, Sumiyo; Ushiki, Aki; Matsuzaki, Hitomi; Ishida, Junji; Sugiyama, Fumihiro; Fukamizu, Akiyoshi

    2014-10-01

    Renin is predominantly expressed in juxtaglomerular cells in the kidney and regulates blood pressure homeostasis. To examine possible in vivo functions of a mouse distal enhancer (mdE), we generated transgenic mice (TgM) carrying either wild-type or mdE-deficient renin BACs (bacterial artificial chromosome), integrated at the identical chromosomal site. In the kidneys of the TgM, the mdE contributed 80% to basal renin promoter activity. To test for possible physiological roles for the mdE, renin BAC transgenes were used to rescue the hypotensive renin-null mice. Interestingly, renal renin expression in the Tg(BAC):renin-null compound mice was indistinguishable between the wild-type and mutant BAC carriers. Surprisingly, however, the plasma renin activity and angiotensin I concentration in the mdE compound mutant mice were significantly lower than the same parameters in the control mice, and the mutants were consistently hypotensive, demonstrating that blood pressure homeostasis is regulated through transcriptional cis elements controlling renin activity.

  15. Sex-specific dysregulation of cysteine oxidation and the methionine and folate cycles in female cystathionine gamma-lyase null mice: a serendipitous model of the methylfolate trap.

    PubMed

    Jiang, Hua; Hurt, K Joseph; Breen, Kelsey; Stabler, Sally P; Allen, Robert H; Orlicky, David J; Maclean, Kenneth N

    2015-08-14

    In addition to its role in the endogenous synthesis of cysteine, cystathionine gamma-lyase (CGL) is a major physiological source of the vasorelaxant hydrogen sulfide. Cgl null mice are potentially useful for studying the influence of this compound upon vascular tone and endothelial function. Here, we confirm a previous report that female Cgl null mice exhibit an approximate 45-fold increase in plasma total homocysteine compared to wild type controls. This level of homocysteine is approximately 3.5-fold higher than that observed in male Cgl null mice and is essentially equivalent to that observed in mouse models of cystathionine beta synthase deficient homocystinuria. Cgl null mice of both sexes exhibited decreased expression of methylenetetrahydrofolate reductase and cysteinesulfinate decarboxylase compared to WT controls. Female Cgl null mice exhibited a sex-specific induction of betaine homocysteine S-methyltransferase and methionine adenosyltransferase 1, alpha and a 70% decrease in methionine synthase expression accompanied by significantly decreased plasma methionine. Decreased plasma cysteine levels in female Cgl null mice were associated with sex-specific dysregulation of cysteine dioxygenase expression. Comparative histological assessment between cystathionine beta-synthase and Cgl null mice indicated that the therapeutic potential of cystathionine against liver injury merits possible further investigation. Collectively, our data demonstrates the importance of considering sex when investigating mouse models of inborn errors of metabolism and indicate that while female Cgl null mice are of questionable utility for studying the physiological role of hydrogen sulfide, they could serve as a useful model for studying the consequences of methionine synthase deficiency and the methylfolate trap.

  16. Sex-specific dysregulation of cysteine oxidation and the methionine and folate cycles in female cystathionine gamma-lyase null mice: a serendipitous model of the methylfolate trap

    PubMed Central

    Jiang, Hua; Hurt, K. Joseph; Breen, Kelsey; Stabler, Sally P.; Allen, Robert H.; Orlicky, David J.; Maclean, Kenneth N.

    2015-01-01

    ABSTRACT In addition to its role in the endogenous synthesis of cysteine, cystathionine gamma-lyase (CGL) is a major physiological source of the vasorelaxant hydrogen sulfide. Cgl null mice are potentially useful for studying the influence of this compound upon vascular tone and endothelial function. Here, we confirm a previous report that female Cgl null mice exhibit an approximate 45-fold increase in plasma total homocysteine compared to wild type controls. This level of homocysteine is approximately 3.5-fold higher than that observed in male Cgl null mice and is essentially equivalent to that observed in mouse models of cystathionine beta synthase deficient homocystinuria. Cgl null mice of both sexes exhibited decreased expression of methylenetetrahydrofolate reductase and cysteinesulfinate decarboxylase compared to WT controls. Female Cgl null mice exhibited a sex-specific induction of betaine homocysteine S-methyltransferase and methionine adenosyltransferase 1, alpha and a 70% decrease in methionine synthase expression accompanied by significantly decreased plasma methionine. Decreased plasma cysteine levels in female Cgl null mice were associated with sex-specific dysregulation of cysteine dioxygenase expression. Comparative histological assessment between cystathionine beta-synthase and Cgl null mice indicated that the therapeutic potential of cystathionine against liver injury merits possible further investigation. Collectively, our data demonstrates the importance of considering sex when investigating mouse models of inborn errors of metabolism and indicate that while female Cgl null mice are of questionable utility for studying the physiological role of hydrogen sulfide, they could serve as a useful model for studying the consequences of methionine synthase deficiency and the methylfolate trap. PMID:26276101

  17. Disruption of Npr1 gene differentially regulates the juxtaglomerular and distal tubular renin levels in null mutant mice

    PubMed Central

    Prieto, Minolfa C; Das, Subhankar; Somanna, Naveen K; Harrison-Bernard, Lisa M; Navar, L Gabriel; Pandey, Kailash N

    2012-01-01

    Atrial natriuretic peptide (ANP) exerts an inhibitory effect on juxtaglomerular (JG) renin synthesis and release by activating guanylyl cyclase/ natriuretic peptide receptor-A (GC-A/NPRA). Renin has also been localized in connecting tubule cells; however, the effect of ANP/NPRA signaling on tubular renin has not been determined. In the present study, we determined the role of NPRA in regulating both JG and tubular renin using Npr1 (coding for NPRA) gene-disrupted mice, which exhibit a hypertensive phenotype. Renin-positive immunoreactivity in Npr1-/- homozygous null mutant mice was significantly reduced compared with Npr1+/+ wild-type mice (23% vs 69% renin-positive glomeruli). However, after chronic diuretic treatment, Npr1-/- mice showed an increment of JG renin immunoreactivity compared with Npr1+/+ mice (70% vs 81% renin-positive glomeruli). There were no significant differences in the distal tubule renin between Npr1+/+ and Npr1-/- mice. However, after diuretic treatment, Npr1-/- mice showed a significant decrease in renin immunoreactivity in principal cells of cortical collecting ducts (p<0.05). The increased JG renin immunoreactivity after reduction in blood pressure in diuretic-treated Npr1-/- mice, demonstrates an inhibitory action of ANP/NPRA system on JG renin; however, a decreased expression of distal tubular renin suggests a differential effect of ANP/NPRA signaling on JG and distal tubular renin. PMID:23071870

  18. Dioxin-dependent and dioxin-independent gene batteries: comparison of liver and kidney in AHR-null mice.

    PubMed

    Boutros, Paul C; Bielefeld, Kirsten A; Pohjanvirta, Raimo; Harper, Patricia A

    2009-11-01

    The aryl hydrocarbon receptor (AHR) is a widely expressed ligand-dependent transcription factor that mediates cellular responses to dioxins and other planar aromatic hydrocarbons. Ahr-null mice are refractory to the toxic effects of dioxin exposure. Although some mechanistic aspects of AHR activity are well understood, the tissue specificity of AHR effects remains unclear, both during development and following administration of exogenous ligands. To address the latter issue, we defined and compared transcriptional responses to dioxin exposure in the liver and kidney of wild-type and Ahr-null adult C57BL/6J mice treated with either 2,3,7,8-tetrachlorodibenzo-p-dioxin or corn-oil vehicle. In both tissues, essentially all effects of dioxin on hepatic mRNA levels were mediated by the AHR. Although 297 genes were altered by dioxin exposure in the liver, only 17 were changed in the kidney, including a number of well-established AHR target genes. Ahr genotype had a large effect in both tissues, profoundly remodeling both the renal and hepatic transcriptomes. Surprisingly, a large number of genes were affected by Ahr genotype in both tissues, suggesting the presence of a basal AHR gene battery. Alterations of the renal transcriptome in Ahr-null animals were associated with perturbation of specific functional pathways and enrichment of specific DNA motifs. Our results demonstrate the importance of intertissue comparisons, highlight the basal role of the AHR in liver and kidney, and support a role in development or normal physiology.

  19. Icariin improves eNOS/NO pathway to prohibit the atherogenesis of apolipoprotein E-null mice.

    PubMed

    Xiao, Hong-Bo; Sui, Guo-Guang; Lu, Xiang-Yang

    2016-12-22

    Impaired endothelial nitric oxide synthase (eNOS)/nitric oxide (NO) pathway induces atherogenesis. The present study examined whether icariin improves the eNOS/NO pathway to prohibit the atherogenesis of apolipoprotein E-null (ApoE(-/-)) mice. In vitro, primary human umbilical vein endothelial cells (HUVECs) were randomly divided into 7 groups: control; vehicle; icariin 10; lyphosphatidylcholine (LPC) group; LPC + icariin 1; LPC + icariin 3; and LPC + icariin 10. In vivo, 80 mice were separated randomly into 4 groups (n = 20): control, ApoE(-/-), ApoE(-/-) + icariin 10, and ApoE(-/-) + icariin 30. ApoE(-/-) mice had significantly more atherosclerosis in the aortic root together with increased aortic ROS production, body mass, plasma triglyceride (TG) and total cholesterol (TC) concentration, decreased aortic eNOS expression, and plasma NO concentration. LPC (10 μg/mL) treatment induced a big decline in NO level in the conditioned medium and eNOS expression, and an increase in intracellular reactive oxygen species (ROS) production in HUVECs. Icariin treatment decreased atherogenesis, ROS production, body mass, plasma TG concentration, and plasma TC concentration, and increased NO concentration and eNOS expression. These findings suggested icariin could improve eNOS/NO-pathway to prohibit the atherogenesis of apolipoprotein E-null mice by restraining oxidative stress.

  20. Reduction of aberrant NF-κB signalling ameliorates Rett syndrome phenotypes in Mecp2-null mice

    PubMed Central

    Kishi, Noriyuki; MacDonald, Jessica L.; Ye, Julia; Molyneaux, Bradley J.; Azim, Eiman; Macklis, Jeffrey D.

    2016-01-01

    Mutations in the transcriptional regulator Mecp2 cause the severe X-linked neurodevelopmental disorder Rett syndrome (RTT). In this study, we investigate genes that function downstream of MeCP2 in cerebral cortex circuitry, and identify upregulation of Irak1, a central component of the NF-κB pathway. We show that overexpression of Irak1 mimics the reduced dendritic complexity of Mecp2-null cortical callosal projection neurons (CPN), and that NF-κB signalling is upregulated in the cortex with Mecp2 loss-of-function. Strikingly, we find that genetically reducing NF-κB signalling in Mecp2-null mice not only ameliorates CPN dendritic complexity but also substantially extends their normally shortened lifespan, indicating broader roles for NF-κB signalling in RTT pathogenesis. These results provide new insight into both the fundamental neurobiology of RTT, and potential therapeutic strategies via NF-κB pathway modulation. PMID:26821816

  1. iNOS null MRL+/+ mice show attenuation of trichloroethene-mediated autoimmunity: contribution of reactive nitrogen species and lipid-derived reactive aldehydes

    PubMed Central

    Wang, Gangduo; Wakamiya, Maki; Wang, Jianling; Ansari, G.A.S.; Khan, M. Firoze

    2015-01-01

    Earlier studies from our laboratory in MRL+/+ mice suggest that free radicals, especially overproduction of reactive nitrogen species (RNS) and lipid-derived reactive aldehydes (LDRAs), are associated with trichloroethene (TCE)-mediated autoimmune response. The current study was undertaken to further assess the contribution of RNS and LDRAs in TCE-mediated autoimmunity by using iNOS-null MRL+/+ mice. iNOS-null MRL+/+ mice were obtained by backcrossing iNOS-null mice (B6.129P2-Nos2tm1Lau/J) to MRL +/+ mice. Female MRL+/+ and iNOS-null MRL+/+ mice were given TCE (10 mmol/kg, i.p., every 4th day) for 6 weeks; their respective controls received corn oil only. TCE exposure led to significantly increased iNOS mRNA in livers, iNOS protein in livers and sera, increased nitrotyrosine (NT) formation in both livers and sera, induction of MDA-/HNE-protein adducts in livers and their respective antibodies in sera along with significant increases in serum antinuclear antibodies (ANA) and anti-dsDNA in MRL+/+ mice. Even though in iNOS-null MRL+/+ mice, the iNOS and NT levels were negligible in both TCE-treated and untreated groups, TCE treatment still led to significant increases in MDA-/HNE-protein adducts and their respective antibodies along with increases in serum ANA and anti-dsDNA compared to controls. Most remarkably, the increases in serum ANA and anti-dsDNA induced by TCE in the iNOS-null MRL+/+ mice were significantly less pronounced compared to that in MRL+/+ mice. Our results provide further evidence that both RNS and LDRAs contribute to TCE-induced autoimmunity in MRL+/+ mice, and iNOS deficiency attenuates this autoimmune response. PMID:26472195

  2. iNOS null MRL+/+ mice show attenuation of trichloroethene-mediated autoimmunity: contribution of reactive nitrogen species and lipid-derived reactive aldehydes.

    PubMed

    Wang, Gangduo; Wakamiya, Maki; Wang, Jianling; Ansari, G A S; Firoze Khan, M

    2015-12-01

    Earlier studies from our laboratory in MRL+/+ mice suggest that free radicals, especially overproduction of reactive nitrogen species (RNS) and lipid-derived reactive aldehydes (LDRAs), are associated with trichloroethene (TCE)-mediated autoimmune response. The current study was undertaken to further assess the contribution of RNS and LDRAs in TCE-mediated autoimmunity by using iNOS-null MRL+/+ mice. iNOS-null MRL+/+ mice were obtained by backcrossing iNOS-null mice (B6.129P2-Nos2(tm1Lau)/J) to MRL +/+ mice. Female MRL+/+ and iNOS-null MRL+/+ mice were given TCE (10 mmol/kg, i.p., every 4(th) day) for 6 weeks; their respective controls received corn oil only. TCE exposure led to significantly increased iNOS mRNA in livers, iNOS protein in livers and sera, increased nitrotyrosine (NT) formation in both livers and sera, induction of MDA-/HNE-protein adducts in livers and their respective antibodies in sera along with significant increases in serum antinuclear antibodies (ANA) and anti-dsDNA in MRL+/+ mice. Even though in iNOS-null MRL+/+ mice, the iNOS and NT levels were negligible in both TCE-treated and untreated groups, TCE treatment still led to significant increases in MDA-/HNE-protein adducts and their respective antibodies along with increases in serum ANA and anti-dsDNA compared to controls. Most remarkably, the increases in serum ANA and anti-dsDNA induced by TCE in the iNOS-null MRL+/+ mice were significantly less pronounced compared to that in MRL+/+ mice. Our results provide further evidence that both RNS and LDRAs contribute to TCE-induced autoimmunity in MRL+/+ mice, and iNOS deficiency attenuates this autoimmune response.

  3. Electrocardiographic and other cardiac anomalies in beta-glucuronidase-null mice corrected by nonablative neonatal marrow transplantation.

    PubMed

    Schuldt, A J T; Hampton, T J; Chu, V; Vogler, C A; Galvin, N; Lessard, M D; Barker, J E

    2004-01-13

    Cardiovascular manifestations of lysosomal storage disease (LSD) are a significant health problem for affected patients. Infantile-onset cardiac disease, because of its rapid progression, is usually treated symptomatically. Therapy in older patients includes valve replacement and bone marrow (BM) transplantation, both of which are life threatening in the already debilitated patients. Enzyme replacement therapy has potential benefit but has not yet been demonstrated to provide long-term relief for cardiac disease. Here, we demonstrate prevention of severe cardiac manifestations in beta-glucuronidase (GUSB) null mice BM-transplanted i.v. as neonates without myeloablative pretreatment. The mice, a model of mucopolysaccharidosis type VII (MPSVII, Sly syndrome), develop progressive LSD unless provided with GUSB early in life. The BM recipients retained GUSB+ donor cells in the peripheral blood and heart until necropsy at > or = 11 months of age. The enzyme beta-hexosamindase increased in tissues of GUSB null MPSVII mice was reduced significantly (P = 0.001) in treated MPSVII hearts. Electrocardiography demonstrated normalization of heart rate, PR, PQ, and QRS intervals in BM recipients. Storage was markedly reduced in the stroma of heart valves, adventitial cells of the aortic root, perivascular and interstitial cells of the myocardium, and interstitial cells of the conduction tissue. Heart/body weight ratio normalized. The aortic root was still grossly distended, and the conductive myocytes retained storage, suggesting neither plays a major role in ECG normalization. We conclude that transplantation of MPSVII neonates without toxic intervention can prevent many of the cardiovascular manifestations of LSD.

  4. Lovastatin fails to improve motor performance and survival in methyl-CpG-binding protein2-null mice

    PubMed Central

    Villani, Claudia; Sacchetti, Giuseppina; Bagnati, Renzo; Passoni, Alice; Fusco, Federica; Carli, Mirjana; Invernizzi, Roberto William

    2016-01-01

    Previous studies provided evidence for the alteration of brain cholesterol homeostasis in 129.Mecp2-null mice, an experimental model of Rett syndrome. The efficacy of statins in improving motor symptoms and prolonging survival of mutant mice suggested a potential role of statins in the therapy of Rett syndrome. In the present study, we show that Mecp2 deletion had no effect on brain and reduced serum cholesterol levels and lovastatin (1.5 mg/kg, twice weekly as in the previous study) had no effects on motor deficits and survival when Mecp2 deletion was expressed on a background strain (C57BL/6J; B6) differing from that used in the earlier study. These findings indicate that the effects of statins may be background specific and raise important issues to consider when contemplating clinical trials. The reduction of the brain cholesterol metabolite 24S-hydroxycholesterol (24S-OHC) found in B6.Mecp2-null mice suggests the occurrence of changes in brain cholesterol metabolism and the potential utility of using plasma levels of 24S-OHC as a biomarker of brain cholesterol homeostasis in RTT. DOI: http://dx.doi.org/10.7554/eLife.22409.001 PMID:27892851

  5. Genetic Disruption of Myostatin Reduces the Development of Proatherogenic Dyslipidemia and Atherogenic Lesions In Ldlr Null Mice

    PubMed Central

    Tu, Powen; Bhasin, Shalender; Hruz, Paul W.; Herbst, Karen L.; Castellani, Lawrence W.; Hua, Ning; Hamilton, James A.; Guo, Wen

    2009-01-01

    OBJECTIVE Insulin-resistant states, such as obesity and type 2 diabetes, contribute substantially to accelerated atherogenesis. Null mutations of myostatin (Mstn) are associated with increased muscle mass and decreased fat mass. In this study, we determined whether Mstn disruption could prevent the development of insulin resistance, proatherogenic dyslipidemia, and atherogenesis. RESEARCH DESIGN AND METHODS C57BL/6 Ldlr−/− mice were cross-bred with C57BL/6 Mstn−/− mice for >10 generations to generate Mstn−/−/Ldlr−/− double-knockout mice. The effects of high-fat/high-cholesterol diet on body composition, plasma lipids, systemic and tissue-specific insulin sensitivity, hepatic steatosis, as well as aortic atheromatous lesion were characterized in Mstn−/−/Ldlr−/− mice in comparison with control Mstn+/+/Ldlr−/− mice. RESULTS Compared with Mstn+/+/Ldlr−/− controls, Mstn−/−/ Ldlr−/− mice were resistant to diet-induced obesity, and had greatly improved insulin sensitivity, as indicated by 42% higher glucose infusion rate and 90% greater muscle [3H]-2-deoxyglucose uptake during hyperinsulinemic-euglycemic clamp. Mstn−/−/Ldlr−/− mice were protected against diet-induced hepatic steatosis and had 56% higher rate of hepatic fatty acid β-oxidation than controls. Mstn−/−/Ldlr−/− mice also had 36% lower VLDL secretion rate and were protected against diet-induced dyslipidemia, as indicated by 30–60% lower VLDL and LDL cholesterol, free fatty acids, and triglycerides. Magnetic resonance angiography and en face analyses demonstrated 41% reduction in aortic atheromatous lesions in Ldlr−/− mice with Mstn deletion. CONCLUSIONS Inactivation of Mstn protects against the development of insulin resistance, proatherogenic dyslipidemia, and aortic atherogenesis in Ldlr−/− mice. Myostatin may be a useful target for drug development for prevention and treatment of obesity and its associated type 2 diabetes and atherosclerosis

  6. Modeling the Cell Muscle Membrane from Normal and Desmin- or Dystrophin-null Mice as an Elastic System

    NASA Astrophysics Data System (ADS)

    García-Pelagio, Karla P.; Santamaría-Holek, Ivan; Bloch, Robert J.; Ortega, Alicia; González-Serratos, Hugo

    2010-12-01

    Two of the most important proteins linking the contractile apparatus and costameres at the sarcolemma of skeletal muscle fibers are dystrophin and desmin. We have developed an elastic model of the proteins that link the sarcolemma to the myofibrils. This is a distributed model, with an elastic constant, k, that includes the main protein components of the costameres. The distributed spring model is composed of parallel units attached in series. To test the model, we performed experiments in which we applied negative pressure, generated by an elastimeter, to a small area of the sarcolemma from single myofiber. The negative pressure formed a bleb of variable height, dependent on the pressure applied. We normalized our measurements of k in dystrophin-null (mdx) and desmin-null (des-/-) mice to the value we obtained for wild type (WT) mice, which was set at 1.0. The relative experimental value for the stiffness of myofibers from mice lacking dystrophin or desmin was 0.5 and 0.7, respectively. The theoretical k values of the individual elements were obtained using neural networks (NN), in which the input was the k value for each parallel spring component and the output was the solution of each resulting parallel system. We compare the experimental values of k in control and mutant muscles to the theoretical values obtained by NN for each protein. Computed theoretical values were 0.4 and 0.8 for dystrophin- and desmin-null muscles, respectively, and 0.9 for WT, in reasonable agreement with our experimental results. This suggests that, although it is a simplified spring model solved by NN, it provides a good approximation of the distribution of spring elements and the elastic constants of the proteins that form the costameres. Our results show that dystrophin is the protein that contributes more than any other to the strength of the connections between the sarcolemma and the contractile apparatus, the costameres.

  7. Enhancement of postsynaptic GABAA and extrasynaptic NMDA receptor-mediated responses in the barrel cortex of Mecp2-null mice

    PubMed Central

    Lo, Fu-Sun; Blue, Mary E.

    2015-01-01

    Rett syndrome (RTT) is a neurodevelopmental disorder that results from mutations in the X-linked gene for methyl-CpG-binding protein 2 (MECP2). The underlying cellular mechanism for the sensory deficits in patients with RTT is largely unknown. This study used the Bird mouse model of RTT to investigate sensory thalamocortical synaptic transmission in the barrel cortex of Mecp2-null mice. Electrophysiological results showed an excitation/inhibition imbalance, biased toward inhibition, due to an increase in efficacy of postsynaptic GABAA receptors rather than alterations in inhibitory network and presynaptic release properties. Enhanced inhibition impaired the transmission of tonic sensory signals from the thalamus to the somatosensory cortex. Previous morphological studies showed an upregulation of NMDA receptors in the neocortex of both RTT patients and Mecp2-null mice at early ages [Blue ME, Naidu S, Johnston MV. Ann Neurol 45: 541–545, 1999; Blue ME, Kaufmann WE, Bressler J, Eyring C, O'Driscoll C, Naidu S, Johnston MV. Anat Rec (Hoboken) 294: 1624–1634, 2011]. Although AMPA and NMDA receptor-mediated excitatory synaptic transmission was not altered in the barrel cortex of Mecp2-null mice, extrasynaptic NMDA receptor-mediated responses increased markedly. These responses were blocked by memantine, suggesting that extrasynaptic NMDA receptors play an important role in the pathogenesis of RTT. The results suggest that enhancement of postsynaptic GABAA and extrasynaptic NMDA receptor-mediated responses may underlie impaired somatosensation and that pharmacological blockade of extrasynaptic NMDA receptors may have therapeutic value for RTT. PMID:26683074

  8. Enhancement of postsynaptic GABAA and extrasynaptic NMDA receptor-mediated responses in the barrel cortex of Mecp2-null mice.

    PubMed

    Lo, Fu-Sun; Blue, Mary E; Erzurumlu, Reha S

    2016-03-01

    Rett syndrome (RTT) is a neurodevelopmental disorder that results from mutations in the X-linked gene for methyl-CpG-binding protein 2 (MECP2). The underlying cellular mechanism for the sensory deficits in patients with RTT is largely unknown. This study used the Bird mouse model of RTT to investigate sensory thalamocortical synaptic transmission in the barrel cortex of Mecp2-null mice. Electrophysiological results showed an excitation/inhibition imbalance, biased toward inhibition, due to an increase in efficacy of postsynaptic GABAA receptors rather than alterations in inhibitory network and presynaptic release properties. Enhanced inhibition impaired the transmission of tonic sensory signals from the thalamus to the somatosensory cortex. Previous morphological studies showed an upregulation of NMDA receptors in the neocortex of both RTT patients and Mecp2-null mice at early ages [Blue ME, Naidu S, Johnston MV. Ann Neurol 45: 541-545, 1999; Blue ME, Kaufmann WE, Bressler J, Eyring C, O'Driscoll C, Naidu S, Johnston MV. Anat Rec (Hoboken) 294: 1624-1634, 2011]. Although AMPA and NMDA receptor-mediated excitatory synaptic transmission was not altered in the barrel cortex of Mecp2-null mice, extrasynaptic NMDA receptor-mediated responses increased markedly. These responses were blocked by memantine, suggesting that extrasynaptic NMDA receptors play an important role in the pathogenesis of RTT. The results suggest that enhancement of postsynaptic GABAA and extrasynaptic NMDA receptor-mediated responses may underlie impaired somatosensation and that pharmacological blockade of extrasynaptic NMDA receptors may have therapeutic value for RTT.

  9. Original Research: ACE2 activator associated with physical exercise potentiates the reduction of pulmonary fibrosis.

    PubMed

    Prata, Luana O; Rodrigues, Carolina R; Martins, Jéssica M; Vasconcelos, Paula C; Oliveira, Fabrício Marcus S; Ferreira, Anderson J; Rodrigues-Machado, Maria da Glória; Caliari, Marcelo V

    2017-01-01

    The interstitial lung diseases are poorly understood and there are currently no studies evaluating the association of physical exercise with an ACE2 activator (DIZE) as a possible treatment for this group of diseases. We evaluate the effects of pharmacological treatment with an angiotensin-converting enzyme 2 activator drug, associated with exercise, on the pulmonary lesions induced by bleomycin. From the 96 male Balb/c mice used in the experiment, only 49 received 8 U/kg of bleomycin (BLM, intratracheally). The mice were divided into control (C) and bleomycin (BLM) groups, sedentary and trained (C-SED, C-EXE, BLM-SED, BLM-EXE), control and bleomycin and also sedentary and trained treated with diminazene (C-SED/E, C-EXE/E, BLM-SED/E, BLM-EXE/E). The animals were trained five days/week, 1 h/day with 60% of the maximum load obtained in a functional capacity test, for four weeks. Diminazene groups were treated (1 mg/kg, by gavage) daily until the end of the experiment. The lungs were collected 48 h after the training program, set in buffered formalin and investigated by Gomori's trichrome, immunohistochemistry of collagen type I, TGF-β1, beta-prolyl-4-hydroxylase, MMP-1 and -2. The BLM-EXE/E group obtained a significant increase in functional capacity, reduced amount of fibrosis and type I collagen, decreased expression of TGF-β1 and beta-prolyl-4-hydroxylase and an increase of metalloproteinase -1, -2 when compared with the other groups. The present research shows, for the first time, that exercise training associated with the activation of ACE2 potentially reduces pulmonary fibrosis.

  10. Aberrant Collagen Composition of the Trabecular Meshwork Results in Reduced Aqueous Humor Drainage and Elevated IOP in MMP-9 Null Mice

    PubMed Central

    De Groef, Lies; Andries, Lien; Siwakoti, Anuja; Geeraerts, Emiel; Bollaerts, Ilse; Noterdaeme, Lut; Etienne, Isabelle; Papageorgiou, Anna-Pia; Stalmans, Ingeborg; Billen, Johan; West-Mays, Judith A.; Moons, Lieve

    2016-01-01

    Purpose Homeostatic turnover of the trabecular meshwork extracellular matrix (ECM) is essential to regulate aqueous humor outflow and to maintain intraocular pressure homeostasis. In this study, we evaluated aqueous humor turnover, intraocular pressure, and trabecular meshwork organization in MMP-9 null mice. Methods Intraocular pressure and aqueous humor turnover were measured in MMP-9 null versus wild-type mice. Morphology of the anterior segment of the eye, with special attention to the structural organization of the trabecular meshwork, was investigated by means of optical coherence tomography, light microscopy, and transmission electron microscopy. Furthermore, using quantitative real-time polymerase chain reaction and immunostainings, we evaluated the ECM composition of the trabecular meshwork. Finally, the integrity and function of the retina and optic nerve were assessed, via optical coherence tomography, histologic techniques, and optomotor testing. Results MMP-9 null mice displayed early-onset ocular hypertension and reduced aqueous humor turnover. While transmission electron microscopic analysis did not reveal any abnormalities in the cellular organization of the trabecular meshwork, detailed investigation of collagen expression indicated that there is an aberrant trabecular meshwork ECM composition in MMP-9 null mice. Notably, at the age of 13 months, no glaucomatous neurodegeneration was seen in MMP-9 null mice. Conclusions Our observations corroborate MMP-9 as an important remodeler of the collagenous composition of the trabecular meshwork and provide evidence for a causal link between MMP-9 deficiency, trabecular meshwork ultrastructure, and ocular hypertension. PMID:27820954

  11. Dioxin-Dependent and Dioxin-Independent Gene Batteries: Comparison of Liver and Kidney in AHR-Null Mice

    PubMed Central

    Boutros, Paul C.; Bielefeld, Kirsten A.; Pohjanvirta, Raimo; Harper, Patricia A.

    2009-01-01

    The aryl hydrocarbon receptor (AHR) is a widely expressed ligand-dependent transcription factor that mediates cellular responses to dioxins and other planar aromatic hydrocarbons. Ahr-null mice are refractory to the toxic effects of dioxin exposure. Although some mechanistic aspects of AHR activity are well understood, the tissue specificity of AHR effects remains unclear, both during development and following administration of exogenous ligands. To address the latter issue, we defined and compared transcriptional responses to dioxin exposure in the liver and kidney of wild-type and Ahr-null adult C57BL/6J mice treated with either 2,3,7,8-tetrachlorodibenzo-p-dioxin or corn-oil vehicle. In both tissues, essentially all effects of dioxin on hepatic mRNA levels were mediated by the AHR. Although 297 genes were altered by dioxin exposure in the liver, only 17 were changed in the kidney, including a number of well-established AHR target genes. Ahr genotype had a large effect in both tissues, profoundly remodeling both the renal and hepatic transcriptomes. Surprisingly, a large number of genes were affected by Ahr genotype in both tissues, suggesting the presence of a basal AHR gene battery. Alterations of the renal transcriptome in Ahr-null animals were associated with perturbation of specific functional pathways and enrichment of specific DNA motifs. Our results demonstrate the importance of intertissue comparisons, highlight the basal role of the AHR in liver and kidney, and support a role in development or normal physiology. PMID:19759094

  12. Pharmacological treatment with mirtazapine rescues cortical atrophy and respiratory deficits in MeCP2 null mice

    PubMed Central

    Bittolo, Tamara; Raminelli, Carlo Antonio; Deiana, Chiara; Baj, Gabriele; Vaghi, Valentina; Ferrazzo, Sara; Bernareggi, Annalisa; Tongiorgi, Enrico

    2016-01-01

    Loss of MeCP2 (Methyl CpG binding protein 2) in Rett syndrome (RTT) causes brain weight decrease, shrinkage of the cortex with reduced dendritic arborization, behavioral abnormalities, seizures and cardio-respiratory complications. The observed monoamine neurotransmitters reduction in RTT suggested antidepressants as a possible therapy. We treated MeCP2-null mice from postnatal-day 28 for two weeks with desipramine, already tested in RTT, or mirtazapine, an antidepressant with limited side-effects, known to promote GABA release. Mirtazapine was more effective than desipramine in restoring somatosensory cortex thickness by fully rescuing pyramidal neurons dendritic arborization and spine density. Functionally, mirtazapine treatment normalized heart rate, breath rate, anxiety levels, and eliminated the hopping behavior observed in MeCP2-null mice, leading to improved phenotypic score. These morphological and functional effects of mirtazapine were accompanied by reestablishment of the GABAergic and glutamatergic receptor activity recorded in cortex and brainstem tissues. Thus, mirtazapine can represent a new potential pharmacological treatment for the Rett syndrome. PMID:26806603

  13. Renoprotective Effects of Vitex megapotamica (Spreng.) Moldenke in C57BL/6 LDLr-Null Mice Undergoing High Fat Diet

    PubMed Central

    Araújo, Valdinei de Oliveira; Gasparotto, Francielly Mourão; Pires, Vanessa Aranega; Maciel, Aline Antunes; Ortmann, Caroline Flach; Cardozo Junior, Euclides Lara; Lourenço, Emerson Luiz Botelho; Gasparotto Junior, Arquimedes

    2015-01-01

    Although Vitex megapotamica (Spreng.) Moldenke is used in Brazilian folk medicine as hypolipidemic drug no study has been conducted to evaluate the effects of this species in an experimental model of atherosclerosis. So, the aim of this study was to evaluate the possible renoprotective activity of methanolic extract obtained from Vitex megapotamica (MEVM) using C57BL/6 LDLr-null mice submitted to high fat diet (HFD). MEVM was orally administered at doses of 30, 100, and 300 mg/kg, for three weeks, starting from the 2nd week of HFD. Systolic blood pressure (SBP) and diuretic activity were measured weekly. At the end of experiments the serum lipids, atherogenic index serum (AIS), oxidative stress, and markers of renal function were determined. HFD induced a significant increase in the systolic blood pressure, dyslipidemia, increase in AIS, and lipid peroxidation accompanied by an important reduction in renal function. Treatment with MEVM was able to prevent increase in SBP, total cholesterol, triglycerides, AIS, urea, and creatinine levels in LDLr-null mice. These effects were accompanied by a significant reduction in oxidative stress and renal injury. The data reported here support the potential of Vitex megapotamica as candidate to be an herbal medicine used in cardiovascular or renal diseases. PMID:25788962

  14. Liver specific expression of Cu/ZnSOD extends the lifespan of Sod1 null mice.

    PubMed

    Zhang, Yiqiang; Liu, Yuhong; Walsh, Michael; Bokov, Alex; Ikeno, Yuji; Jang, Young C; Perez, Viviana I; Van Remmen, Holly; Richardson, Arlan

    2016-03-01

    Genetic ablation of CuZn-superoxide dismutase (Sod1) in mice (Sod1(-/-) mice) leads to shortened lifespan with a dramatic increase in hepatocellular carcinoma and accelerated aging phenotypes, including early onset sarcopenia. To study the tissue specific effects of oxidative stress in the Sod1(-/-) mice, we generated mice that only express the human SOD1 gene specifically in the liver of Sod1(-/-) mice (Sod1(-/-)/hSOD1(alb) mice). Expression of hSOD1 in the liver of Sod1(-/-) mice improved liver function, reduced oxidative damage in liver, and partially restored the expression of several genes involved in tumorigenesis, which are abnormally expressed in the livers of the Sod1(-/-) mice. However, liver specific expression of hSOD1 did not prevent the loss of body weight and muscle mass and alterations in the structure of neuromuscular junctions. The expression of hSOD1 in the liver of Sod1(-/-) mice significantly improved the lifespan of Sod1(-/-) mice; however, the lifespan of the Sod1(-/-)/hSOD1(alb) mice was still significantly shorter than wild type mice.

  15. Calmodulin interacts with angiotensin-converting enzyme-2 (ACE2) and inhibits shedding of its ectodomain.

    PubMed

    Lambert, Daniel W; Clarke, Nicola E; Hooper, Nigel M; Turner, Anthony J

    2008-01-23

    Angiotensin-converting enzyme-2 (ACE2) is a regulatory protein of the renin-angiotensin system (RAS) and a receptor for the causative agent of severe-acute respiratory syndrome (SARS), the SARS-coronavirus. We have previously shown that ACE2 can be shed from the cell surface in response to phorbol esters by a process involving TNF-alpha converting enzyme (TACE; ADAM17). In this study, we demonstrate that inhibitors of calmodulin also stimulate shedding of the ACE2 ectodomain, a process at least partially mediated by a metalloproteinase. We also show that calmodulin associates with ACE2 and that this interaction is decreased by calmodulin inhibitors.

  16. Apamin Boosting of Synaptic Potentials in CaV2.3 R-Type Ca2+ Channel Null Mice.

    PubMed

    Wang, Kang; Kelley, Melissa H; Wu, Wendy W; Adelman, John P; Maylie, James

    2015-01-01

    SK2- and KV4.2-containing K+ channels modulate evoked synaptic potentials in CA1 pyramidal neurons. Each is coupled to a distinct Ca2+ source that provides Ca2+-dependent feedback regulation to limit AMPA receptor (AMPAR)- and NMDA receptor (NMDAR)-mediated postsynaptic depolarization. SK2-containing channels are activated by Ca2+ entry through NMDARs, whereas KV4.2-containing channel availability is increased by Ca2+ entry through SNX-482 (SNX) sensitive CaV2.3 R-type Ca2+ channels. Recent studies have challenged the functional coupling between NMDARs and SK2-containing channels, suggesting that synaptic SK2-containing channels are instead activated by Ca2+ entry through R-type Ca2+ channels. Furthermore, SNX has been implicated to have off target affects, which would challenge the proposed coupling between R-type Ca2+ channels and KV4.2-containing K+ channels. To reconcile these conflicting results, we evaluated the effect of SK channel blocker apamin and R-type Ca2+ channel blocker SNX on evoked excitatory postsynaptic potentials (EPSPs) in CA1 pyramidal neurons from CaV2.3 null mice. The results show that in the absence of CaV2.3 channels, apamin application still boosted EPSPs. The boosting effect of CaV2.3 channel blockers on EPSPs observed in neurons from wild type mice was not observed in neurons from CaV2.3 null mice. These data are consistent with a model in which SK2-containing channels are functionally coupled to NMDARs and KV4.2-containing channels to CaV2.3 channels to provide negative feedback regulation of EPSPs in the spines of CA1 pyramidal neurons.

  17. Relationship between hepatic phenotype and changes in gene expression in cytochrome P450 reductase (POR) null mice

    PubMed Central

    Wang, Xiu Jun; Chamberlain, Mark; Vassieva, Olga; Henderson, Colin J.; Wolf, C. Roland

    2005-01-01

    Cytochrome P450 reductase is the unique electron donor for microsomal cytochrome P450s; these enzymes play a major role in the metabolism of endogenous and xenobiotic compounds. In mice with a liver-specific deletion of cytochrome P450 reductase, hepatic cytochrome P450 activity is ablated, with consequent changes in bile acid and lipid homoeostasis. In order to gain insights into the metabolic changes resulting from this phenotype, we have analysed changes in hepatic mRNA expression using microarray analysis and real-time PCR. In parallel with the perturbations in bile acid levels, changes in the expression of key enzymes involved in cholesterol and lipid homoeostasis were observed in hepatic cytochrome P450 reductase null mice. This was characterized by a reduced expression of Cyp7b1, and elevation of Cyp7a1 and Cyp8b1 expression. The levels of mRNAs for other cytochrome P450 genes, including Cyp2b10, Cyp2c29, Cyp3a11 and Cyp3a16, were increased, demonstrating that endogenous factors play a role in regulating the expression of these proteins and that the increases are due, at least in part, to altered levels of transcripts. In addition, levels of mRNAs encoding genes involved in glycolysis and lipid transport were also increased; the latter may provide an explanation for the increased hepatic lipid content observed in the hepatic null mice. Serum testosterone and oestradiol levels were lowered, accompanied by significantly decreased expression of Hsd3b2 (3β-hydroxy-Δ5-steroid dehydrogenase-2), Hsd3b5 (3β-hydroxy-Δ5-steroid dehydrogenase-5) and Hsd11b1 (11β-hydroxysteroid dehydrogenase type 1), key enzymes in steroid hormone metabolism. These microarray data provide important insights into the control of metabolic pathways by the cytochrome system. PMID:15717863

  18. Distribution of mercury in metallothionein-null mice after exposure to mercury vapor: amount of metallothionein isoform does not affect accumulation of mercury in the brain.

    PubMed

    Yasutake, Akira; Yoshida, Minoru; Honda, Akiko; Watanabe, Chiho; Satoh, Masahiko

    2012-01-01

    To examine the contribution of metallothionein (MT) to mercury accumulation in mouse tissues, 129 strain female mice and MT null mice were exposed to metallic mercury vapor at a sub-toxic level, and Hg levels in the brain, kidney and liver were determined on 1, 3 and 7 days after the exposure. After exposure to mercury vapor, significant Hg accumulation was observed in the brains of wild-type and MT-I/II null and MT-III null mice, as well as in the liver and kidneys. No strain difference was observed in the tissue Hg accumulations 24 hr after the exposure except for the kidneys, where the highest accumulation was found in MT-III null mice. Although the brains of MT-III null mice showed slightly higher Hg accumulation than the other two strains, no significant difference was observed except in the cerebrum on Day 7. Gel chromatograms of cerebrum soluble fractions revealed that a significant amount of Hg existed as an MT-bound form in all the mouse strains. On the other hand, MT-bound Hg was found as a minor fraction in soluble fractions of the kidneys and livers in wild-type and MT-III null mice. Despite a significant strain difference in total MT levels in the cerebrum, there was no difference among the three strains in the amount of Hg accumulated in the cerebrum and its distribution rates in MT fractions. The present study demonstrated that brain uptake of Hg(0) and its accumulation as Hg(2+) did not depend on the amount of MT isoform in the tissue, at least in the early phase.

  19. Decreased bile-acid synthesis in livers of hepatocyte-conditional NADPH-cytochrome P450 reductase-null mice results in increased bile acids in serum.

    PubMed

    Cheng, Xingguo; Zhang, Youcai; Klaassen, Curtis D

    2014-10-01

    NADPH-cytochrome P450 reductase (Cpr) is essential for the function of microsomal cytochrome P450 monooxygenases (P450), including those P450s involved in bile acid (BA) synthesis. Mice with hepatocyte-specific deletion of NADPH-cytochrome P450 reductase (H-Cpr-null) have been engineered to understand the in vivo function of hepatic P450s in the metabolism of xenobiotics and endogenous compounds. However, the impact of hepatic Cpr on BA homeostasis is not clear. The present study revealed that H-Cpr-null mice had a 60% decrease in total BA concentration in liver, whereas the total BA concentration in serum was almost doubled. The decreased level of cholic acid (CA) in both serum and livers of H-Cpr-null mice is likely due to diminished enzyme activity of Cyp8b1 that is essential for CA biosynthesis. Feedback mechanisms responsible for the reduced liver BA concentrations and/or increased serum BA concentrations in H-Cpr-null mice included the following: 1) enhanced alternative BA synthesis pathway, as evidenced by the fact that classic BA synthesis is diminished but chenodeoxycholic acid still increases in both serum and livers of H-Cpr-null mice; 2) inhibition of farnesoid X receptor activation, which increased the mRNA of Cyp7a1 and 8b1; 3) induction of intestinal BA transporters to facilitate BA absorption from the intestine to the circulation; 4) induction of hepatic multidrug resistance-associated protein transporters to increase BA efflux from the liver to blood; and 5) increased generation of secondary BAs. In summary, the present study reveals an important contribution of the alternative BA synthesis pathway and BA transporters in regulating BA concentrations in H-Cpr-null mice.

  20. Similar L-dopa-stimulated motor activity in mice with adult-onset 6-hydroxydopamine-induced symmetric dopamine denervation and in transcription factor Pitx3 null mice with perinatal-onset symmetric dopamine denervation.

    PubMed

    Li, Li; Sagot, Ben; Zhou, Fu-Ming

    2015-07-30

    The transcription factor Pitx3 null mutant (Pitx3Null) mice have a constitutive perinatal-onset and symmetric bilateral dopamine (DA) loss in the striatum. In these mice l-3,4-dihydroxyphenylalanine (l-dopa) induces apparently normal horizontal movements (walking) but also upward movements consisting of the vertical body trunk and waving paws that are absent in normal animals and in animals with the classic unilateral 6-hydroxydopamine (6-OHDA) lesion-induced DA denervation. Thus, a concern is that the perinatal timing of the DA loss and potential developmental abnormalities in Pitx3Null mice may underlie these upward movements, thus reducing the usefulness as a DA denervation model. Here we show that in normal wild-type (Pitx3WT) mice with adult-onset symmetric, bilateral 6-OHDA-induced DA lesion in the dorsal striatum, l-dopa induces normal horizontal movements and upward movements that are qualitatively identical to those in Pitx3Null mice. Furthermore, after unilateral 6-OHDA lesion of the residual DA innervation in the striatum in Pitx3Null mice, l-dopa induces contraversive rotation that is similar to that in Pitx3WT mice with the classic unilateral 6-OHDA lesion. These results indicate that in Pitx3Null mice, the bilateral symmetric DA denervation in the dorsal striatum is sufficient for expressing the l-dopa-induced motor phenotype and the perinatal timing of their DA loss is not a determining factor, providing further evidence that Pitx3Null mice are a convenient and suitable mouse model to study the consequences of DA loss and dopaminergic replacement therapy in Parkinson's disease.

  1. Prohormone convertase 2 (PC2) null mice have increased mu opioid receptor levels accompanied by altered morphine-induced antinociception, tolerance and dependence.

    PubMed

    Lutfy, K; Parikh, D; Lee, D L; Liu, Y; Ferrini, M G; Hamid, A; Friedman, T C

    2016-08-04

    Chronic morphine treatment increases the levels of prohormone convertase 2 (PC2) in brain regions involved in nociception, tolerance and dependence. Thus, we tested if PC2 null mice exhibit altered morphine-induced antinociception, tolerance and dependence. PC2 null mice and their wild-type controls were tested for baseline hot plate latency, injected with morphine (1.25-10mg/kg) and tested for antinociception 30min later. For tolerance studies, mice were tested in the hot plate test before and 30min following morphine (5mg/kg) on day 1. Mice then received an additional dose so that the final dose of morphine was 10mg/kg on this day. On days 2-4, mice received additional doses of morphine (20, 40 and 80mg/kg on days 1, 2, 3, and 4, respectively). On day 5, mice were tested in the hot plate test before and 30min following morphine (5mg/kg). For withdrawal studies, mice were treated with the escalating doses of morphine (10, 20, 40 and 80mg/kg) for 4days, implanted with a morphine pellet on day 5 and 3 days later injected with naloxone (1mg/kg) and signs of withdrawal were recorded. Morphine dose-dependently induced antinociception and the magnitude of this response was greater in PC2 null mice. Tolerance to morphine was observed in wild-type mice and this phenomenon was blunted in PC2 null mice. Withdrawal signs were also reduced in PC2 null mice. Immunohistochemical studies showed up-regulation of the mu opioid receptor (MOP) protein expression in the periaqueductal gray area, ventral tegmental area, lateral hypothalamus, medial hypothalamus, nucleus accumbens, and somatosensory cortex in PC2 null mice. Likewise, naloxone specific binding was increased in the brains of these mice compared to their wild-type controls. The results suggest that the PC2-derived peptides may play a functional role in morphine-induced antinociception, tolerance and dependence. Alternatively, lack of opioid peptides led to up-regulation of the MOP and altered morphine

  2. Microglia contribute to circuit defects in Mecp2 null mice independent of microglia-specific loss of Mecp2 expression

    PubMed Central

    Schafer, Dorothy P; Heller, Christopher T; Gunner, Georgia; Heller, Molly; Gordon, Christopher; Hammond, Timothy; Wolf, Yochai; Jung, Steffen; Stevens, Beth

    2016-01-01

    Microglia, the resident CNS macrophages, have been implicated in the pathogenesis of Rett Syndrome (RTT), an X-linked neurodevelopmental disorder. However, the mechanism by which microglia contribute to the disorder is unclear and recent data suggest that microglia do not play a causative role. Here, we use the retinogeniculate system to determine if and how microglia contribute to pathogenesis in a RTT mouse model, the Mecp2 null mouse (Mecp2tm1.1Bird/y). We demonstrate that microglia contribute to pathogenesis by excessively engulfing, thereby eliminating, presynaptic inputs at end stages of disease (≥P56 Mecp2 null mice) concomitant with synapse loss. Furthermore, loss or gain of Mecp2 expression specifically in microglia (Cx3cr1CreER;Mecp2fl/yor Cx3cr1CreER; Mecp2LSL/y) had little effect on excessive engulfment, synapse loss, or phenotypic abnormalities. Taken together, our data suggest that microglia contribute to end stages of disease by dismantling neural circuits rendered vulnerable by loss of Mecp2 in other CNS cell types. DOI: http://dx.doi.org/10.7554/eLife.15224.001 PMID:27458802

  3. Transforming growth factor-beta1 null mutation causes infertility in male mice associated with testosterone deficiency and sexual dysfunction.

    PubMed

    Ingman, Wendy V; Robertson, Sarah A

    2007-08-01

    TGFbeta1 is a multifunctional cytokine implicated in gonad and secondary sex organ development, steroidogenesis, and spermatogenesis. To determine the physiological requirement for TGFbeta1 in male reproduction, Tgfb1 null mutant mice on a Prkdc(scid) immunodeficient background were studied. TGFbeta1-deficient males did not deposit sperm or induce pseudopregnancy in females, despite an intact reproductive tract with morphologically normal penis, seminal vesicles, and testes. Serum and intratesticular testosterone and serum androstenedione were severely diminished in TGFbeta1-deficient males. Testosterone deficiency was secondary to disrupted pituitary gonadotropin secretion because serum LH and to a lesser extent serum FSH were reduced, and exogenous LH replacement with human chorionic gonadotropin (hCG) induced serum testosterone to control levels. In the majority of TGFbeta1-deficient males, spermatogenesis was normal and sperm were developmentally competent as assessed by in vitro fertilization. Analysis of sexual behavior revealed that although TGFbeta1 null males showed avid interest in females and engaged in mounting activity, intromission was infrequent and brief, and ejaculation was not attained. Administration of testosterone to adult males, even after neonatal androgenization, was ineffective in restoring sexual function; however, erectile reflexes and ejaculation could be induced by electrical stimulation. These studies demonstrate the profound effect of genetic deficiency in TGFbeta1 on male fertility, implicating this cytokine in essential roles in the hypothalamic-pituitary-gonadal axis and in testosterone-independent regulation of mating competence.

  4. CMP-Neu5Ac Hydroxylase Null Mice as a Model for Studying Metabolic Disorders Caused by the Evolutionary Loss of Neu5Gc in Humans

    PubMed Central

    Kwon, Deug-Nam; Choi, Yun-Jung; Cho, Ssang-Goo; Park, Chankyu; Seo, Han Geuk; Song, Hyuk; Kim, Jin-Hoi

    2015-01-01

    The purpose of this study was to identify the modification/turnover of gene products that are altered in humans due to evolutionary loss of Neu5Gc. CMP-Neu5Ac hydroxylase- (Cmah-) deficient mice show the infiltration of Kupffer cells within liver sinusoids, whereas body and liver weight develop normally. Pathway analysis by use of Illumina MouseRef-8 v2 Expression BeadChip provided evidence that a number of biological pathways, including the glycolysis, gluconeogenesis, TCA cycle, and pentose phosphate pathways, as well as glycogen metabolism-related gene expression, were significantly upregulated in Cmah-null mice. The intracellular glucose supply in Cmah-null mice resulted in mitochondrial dysfunction, oxidative stress, and the advanced glycation end products accumulation that could further induce oxidative stress. Finally, low sirtuin-1 and sirtuin-3 gene expressions due to higher NADH/NAD in Cmah-null mice decreased Foxo-1 and MnSOD gene expression, suggesting that oxidative stress may result in mitochondrial dysfunction in Cmah-null mouse. The present study suggests that mice with CMAH deficiency can be taken as an important model for studying metabolic disorders in humans. PMID:26558285

  5. Neil2-null Mice Accumulate Oxidized DNA Bases in the Transcriptionally Active Sequences of the Genome and Are Susceptible to Innate Inflammation.

    PubMed

    Chakraborty, Anirban; Wakamiya, Maki; Venkova-Canova, Tatiana; Pandita, Raj K; Aguilera-Aguirre, Leopoldo; Sarker, Altaf H; Singh, Dharmendra Kumar; Hosoki, Koa; Wood, Thomas G; Sharma, Gulshan; Cardenas, Victor; Sarkar, Partha S; Sur, Sanjiv; Pandita, Tej K; Boldogh, Istvan; Hazra, Tapas K

    2015-10-09

    Why mammalian cells possess multiple DNA glycosylases (DGs) with overlapping substrate ranges for repairing oxidatively damaged bases via the base excision repair (BER) pathway is a long-standing question. To determine the biological role of these DGs, null animal models have been generated. Here, we report the generation and characterization of mice lacking Neil2 (Nei-like 2). As in mice deficient in each of the other four oxidized base-specific DGs (OGG1, NTH1, NEIL1, and NEIL3), Neil2-null mice show no overt phenotype. However, middle-aged to old Neil2-null mice show the accumulation of oxidative genomic damage, mostly in the transcribed regions. Immuno-pulldown analysis from wild-type (WT) mouse tissue showed the association of NEIL2 with RNA polymerase II, along with Cockayne syndrome group B protein, TFIIH, and other BER proteins. Chromatin immunoprecipitation analysis from mouse tissue showed co-occupancy of NEIL2 and RNA polymerase II only on the transcribed genes, consistent with our earlier in vitro findings on NEIL2's role in transcription-coupled BER. This study provides the first in vivo evidence of genomic region-specific repair in mammals. Furthermore, telomere loss and genomic instability were observed at a higher frequency in embryonic fibroblasts from Neil2-null mice than from the WT. Moreover, Neil2-null mice are much more responsive to inflammatory agents than WT mice. Taken together, our results underscore the importance of NEIL2 in protecting mammals from the development of various pathologies that are linked to genomic instability and/or inflammation. NEIL2 is thus likely to play an important role in long term genomic maintenance, particularly in long-lived mammals such as humans.

  6. Neil2-null Mice Accumulate Oxidized DNA Bases in the Transcriptionally Active Sequences of the Genome and Are Susceptible to Innate Inflammation* ♦

    PubMed Central

    Chakraborty, Anirban; Wakamiya, Maki; Venkova-Canova, Tatiana; Pandita, Raj K.; Aguilera-Aguirre, Leopoldo; Sarker, Altaf H.; Singh, Dharmendra Kumar; Hosoki, Koa; Wood, Thomas G.; Sharma, Gulshan; Cardenas, Victor; Sarkar, Partha S.; Sur, Sanjiv; Pandita, Tej K.; Boldogh, Istvan; Hazra, Tapas K.

    2015-01-01

    Why mammalian cells possess multiple DNA glycosylases (DGs) with overlapping substrate ranges for repairing oxidatively damaged bases via the base excision repair (BER) pathway is a long-standing question. To determine the biological role of these DGs, null animal models have been generated. Here, we report the generation and characterization of mice lacking Neil2 (Nei-like 2). As in mice deficient in each of the other four oxidized base-specific DGs (OGG1, NTH1, NEIL1, and NEIL3), Neil2-null mice show no overt phenotype. However, middle-aged to old Neil2-null mice show the accumulation of oxidative genomic damage, mostly in the transcribed regions. Immuno-pulldown analysis from wild-type (WT) mouse tissue showed the association of NEIL2 with RNA polymerase II, along with Cockayne syndrome group B protein, TFIIH, and other BER proteins. Chromatin immunoprecipitation analysis from mouse tissue showed co-occupancy of NEIL2 and RNA polymerase II only on the transcribed genes, consistent with our earlier in vitro findings on NEIL2's role in transcription-coupled BER. This study provides the first in vivo evidence of genomic region-specific repair in mammals. Furthermore, telomere loss and genomic instability were observed at a higher frequency in embryonic fibroblasts from Neil2-null mice than from the WT. Moreover, Neil2-null mice are much more responsive to inflammatory agents than WT mice. Taken together, our results underscore the importance of NEIL2 in protecting mammals from the development of various pathologies that are linked to genomic instability and/or inflammation. NEIL2 is thus likely to play an important role in long term genomic maintenance, particularly in long-lived mammals such as humans. PMID:26245904

  7. Cystatin C deficiency increases elastic lamina degradation and aortic dilatation in apolipoprotein E-null mice.

    PubMed

    Sukhova, Galina K; Wang, Bing; Libby, Peter; Pan, Jie-Hong; Zhang, Yaou; Grubb, Anders; Fang, Kenneth; Chapman, Harold A; Shi, Guo-Ping

    2005-02-18

    The pathogenesis of atherosclerosis and abdominal aortic aneurysm involves substantial proteolysis of the arterial extracellular matrix. The lysosomal cysteine proteases can exert potent elastolytic and collagenolytic activity. Human atherosclerotic plaques have increased cysteine protease content and decreased levels of the endogenous inhibitor cystatin C, suggesting an imbalance that would favor matrix degradation in the arterial wall. This study tested directly the hypothesis that impaired expression of cystatin C alters arterial structure. Cystatin C-deficient mice (Cyst C-/-) were crossbred with apolipoprotein E-deficient mice (ApoE-/-) to generate cystatin C and apolipoprotein E-double deficient mice (Cyst C-/-ApoE-/-). After 12 weeks on an atherogenic diet, cystatin C deficiency yielded significantly increased tunica media elastic lamina fragmentation, decreased medial size, and increased smooth muscle cell and collagen content in aortic lesions of ApoE-/- mice. Cyst C-/-ApoE-/- mice also showed dilated thoracic and abdominal aortae compared with control ApoE-/- mice, although atheroma lesion size, intimal macrophage accumulation, and lipid core size did not differ between these mice. These findings demonstrate directly the importance of cysteine protease/protease inhibitor balance in dysregulated arterial integrity and remodeling during experimental atherogenesis.

  8. ACE2 orthologues in non-mammalian vertebrates (Danio, Gallus, Fugu, Tetraodon and Xenopus).

    PubMed

    Chou, Chih-Fong; Loh, Chay Boon; Foo, Yik Khoon; Shen, Shuo; Fielding, Burtram C; Tan, Timothy H P; Khan, Sehaam; Wang, Yue; Lim, Seng Gee; Hong, Wanjin; Tan, Yee-Joo; Fu, Jianlin

    2006-08-01

    Angiotensin-converting enzyme 2 (ACE2), a newly identified member in the renin-angiotensin system (RAS), acts as a negative regulator of ACE. It is mainly expressed in cardiac blood vessels and the tubular epithelia of kidneys and abnormal expression has been implicated in diabetes, hypertension and heart failure. The mechanism and physiological function of this zinc metallopeptidase in mammals are not yet fully understood. Non-mammalian vertebrate models offer attractive and simple alternatives that could facilitate the exploration of ACE2 function. In this paper we report the in silico analysis of Ace2 genes from the Gallus (chicken), Xenopus (frog), Fugu and Tetraodon (pufferfish) genome assembly databases, and from the Danio (zebrafish) cDNA library. Exon ambiguities of Danio and Xenopus Ace2s were resolved by RT-PCR and 3'RACE. Analyses of the exon-intron structures, alignment, phylogeny and hydrophilicity plots, together with the conserved synteny among these vertebrates, support the orthologous relationship between mammalian and non-mammalian ACE2s. The putative promoters of Ace2 from human, Tetraodon and Xenopus tropicalis drove the expression of enhanced green fluorescent protein (EGFP) specifically in the heart tissue of transgenic Xenopus thus making it a suitable model for future functional genomic studies. Additionally, the search for conserved cis-elements resulted in the discovery of WGATAR motifs in all the putative Ace2 promoters from 7 different animals, suggesting a possible role of GATA family transcriptional factors in regulating the expression of Ace2.

  9. Effect of ghrelin receptor antagonist on meal patterns in cholecystokinin type 1 receptor null mice.

    PubMed

    Lee, Jennifer; Martin, Elizabeth; Paulino, Gabriel; de Lartigue, Guillaume; Raybould, Helen E

    2011-05-03

    Vagal afferent neurons (VAN) express the cholecystokinin (CCK) type 1 receptor (CCK₁R) and, as predicted by the role of CCK in inducing satiation, CCK₁R⁻/⁻ mice ingest larger and longer meals. However, after a short fast, CCK₁R⁻/⁻ mice ingesting high fat (HF) diets initiate feeding earlier than wild-type mice. We hypothesized that the increased drive to eat in CCK₁R⁻/⁻ mice eating HF diet is mediated by ghrelin, a gut peptide that stimulates food intake. The decrease in time to first meal, and the increase in meal size and duration in CCK₁R⁻/⁻ compared to wild-type mice ingesting high fat (HF) diet were reversed by administration of GHSR1a antagonist D-(Lys3)-GHRP-6 (p<0.05). Administration of the GHSR1a antagonist significantly increased expression of the neuropeptide cocaine and amphetamine-regulated transcript (CART) in VAN of HF-fed CCK₁R⁻/⁻ but not wild-type mice. Administration of the GHSR1a antagonist decreased neuronal activity measured by immunoreactivity for fos protein in the nucleus of the solitary tract (NTS) and the arcuate nucleus of both HF-fed wild-type and CCK₁R⁻/⁻ mice. The data show that hyperphagia in CCK₁R⁻/⁻ mice ingesting HF diet is reversed by blockade of the ghrelin receptor, suggesting that in the absence of the CCK₁R, there is an increased ghrelin-dependent drive to feed. The site of action of ghrelin receptors is unclear, but may involve an increase in expression of CART peptide in VAN in HF-fed CCK₁R⁻/⁻ mice.

  10. Soluble Epoxide Hydrolase Null Mice Exhibit Female and Male Differences In Regulation of Vascular Homeostasis

    PubMed Central

    Vanella, Luca; Canestraro, Martina; Lee, Craig R.; Cao, Jian; Zeldin, Darryl C.; Schwartzman, Michal L.; Abraham, Nader G.

    2015-01-01

    Increased CYP epoxygenase activity and consequently up regulation of epoxyeicosatrienoic acids (EETs) levels provides protection against metabolic syndrome and cardiovascular diseases. Conversion of arachidonic acid epoxides to diols by soluble epoxide hydrolase (sEH) diminishes the beneficial cardiovascular properties of these epoxyeicosanoids. We therefore examined the possible biochemical consequences of sEH deletion on vascular responses in male and female mice. Through the use of the sEH KO mouse, we provide evidence of differences in the compensatory response in the balance between nitric oxide (NO), carbon monoxide (CO), EETs and the vasoconstrictor 20-HETE in male and female KO mice. Serum levels of adiponectin, TNFα, IL-1b and MCP1 and protein expression in vascular tissue of p-AMPK, p-AKT and p-eNOS were measured. Deletion of sEH caused a significant (p<0,05) decrease in body weight, and an increase in adiponectin, pAMPK and pAKT levels in female KO mice compared to male KO mice. Gene deletion resulted in a higher production of renal EETs in female KO compared to male KO mice and, concomitantly, we observed an increase in renal 20-HETEs levels and superoxide anion production only in male KO mice. sEH deletion increased p-AKT and p-eNOS protein expression but decreased p-AMPK levels in female KO mice. Increased levels of p-eNOS at Thr-495 were observed only in KO male mice. While p-eNOS at 1177 were not significantly different between male and female. Nitric oxide production was unaltered in male KO mice. These results provide evidence of gender differences in the preservation of vascular homeostasis in response to sEH deletion which involves regulation of phosphorylation of eNOS at the 495 site. PMID:25908301

  11. Oxidative stress and ROS metabolism via down-regulation of sirtuin 3 expression in Cmah-null mice affect hearing loss

    PubMed Central

    Choi, Yun-Jung; Gurunathan, Sangiliyandi; Kim, Jin-Hoi

    2015-01-01

    CMP-Neu5Ac hydroxylase (Cmah) disruption caused several abnormalities and diseases including hearing loss in old age. However, underling molecular mechanisms that give rise to age-related hearing loss (AHL) in Cmah-null mouse are still obscure. In this study, Cmah-null mice showed age-related decline of hearing associated with loss of sensory hair cells, spiral ganglion neurons, and/or stria vascularis degeneration in the cochlea. To identify differential gene expression profiles and pathway associated with AHL, we performed microarray analysis using Illumina MouseRef-8 v2 Expression BeadChip and pathway-focused PCR array in the cochlear tissues of Cmah-null mouse. Pathway and molecular mechanism analysis using differentially expressed genes provided evidences that altered biological pathway due to oxidative damage by low expressed antioxidants and dysregulated reactive oxygen species (ROS) metabolism. Especially, low sirtuin 3 (Sirt3) gene expressions in Cmah-null mice decreased both of downstream regulator (Foxo1 and MnSod) and regulatory transcription factor (Hif1α and Foxo3a) gene expression. Taken together, we suggest that down-regulation of Sirt3 expression leads to oxidative stress and mitochondrial dysfunction by regulation of ROS and that it could alter various signaling pathways in Cmah-null mice with AHL. PMID:26319214

  12. Leptin Resistance Contributes to Obesity in Mice with Null Mutation of Carcinoembryonic Antigen-related Cell Adhesion Molecule 1.

    PubMed

    Heinrich, Garrett; Russo, Lucia; Castaneda, Tamara R; Pfeiffer, Verena; Ghadieh, Hilda E; Ghanem, Simona S; Wu, Jieshen; Faulkner, Latrice D; Ergün, Süleyman; McInerney, Marcia F; Hill, Jennifer W; Najjar, Sonia M

    2016-05-20

    Carcinoembryonic antigen-related cell adhesion molecule 1 (CEACAM1) promotes hepatic insulin clearance. Consistently, mice with null mutation of Ceacam1 (Cc1(-/-)) exhibit impaired insulin clearance with increased lipid production in liver and redistribution to white adipose tissue, leading to visceral obesity at 2 months of age. When the mutation is propagated on the C57/BL6J genetic background, total fat mass rises significantly with age, and glucose intolerance and systemic insulin resistance develop at 6 months of age. This study was carried out to determine the mechanisms underlying the marked increase in total fat mass in 6-month-old mutants. Indirect calorimetry analysis showed that Cc1(-/-) mice develop hyperphagia and a significant reduction in physical activity, in particular in the early hours of the dark cycle, during which energy expenditure is only slightly lower than in wild-type mice. They also exhibit increased triglyceride accumulation in skeletal muscle, due in part to incomplete fatty acid β-oxidation. Mechanistically, hypothalamic leptin signaling is reduced, as demonstrated by blunted STAT3 phosphorylation in coronal sections in response to an intracerebral ventricular injection of leptin. Hypothalamic fatty-acid synthase activity is also elevated in the mutants. Together, the data show that the increase in total fat mass in Cc1(-/-) mice is mainly attributed to hyperphagia and reduced spontaneous physical activity. Although the contribution of the loss of CEACAM1 from anorexigenic proopiomelanocortin neurons in the arcuate nucleus is unclear, leptin resistance and elevated hypothalamic fatty-acid synthase activity could underlie altered energy balance in these mice.

  13. Altered Body Weight Regulation in CK1ε Null and tau Mutant Mice on Regular Chow and High Fat Diets

    PubMed Central

    Zhou, Lili; Summa, Keith C.; Olker, Christopher; Vitaterna, Martha H.; Turek, Fred W.

    2016-01-01

    Disruption of circadian rhythms results in metabolic dysfunction. Casein kinase 1 epsilon (CK1ε) is a canonical circadian clock gene. Null and tau mutations in CK1ε show distinct effects on circadian period. To investigate the role of CK1ε in body weight regulation under both regular chow (RC) and high fat (HF) diet conditions, we examined body weight on both RC and HF diets in CK1ε−/− and CK1εtau/tau mice on a standard 24 hr light-dark (LD) cycle. Given the abnormal entrainment of CK1εtau/tau mice on a 24 hr LD cycle, a separate set of CK1εtau/tau mice were tested under both diet conditions on a 20 hr LD cycle, which more closely matches their endogenous period length. On the RC diet, both CK1ε−/− and CK1εtau/tau mutants on a 24 hr LD cycle and CK1εtau/tau mice on a 20 hr LD cycle exhibited significantly lower body weights, despite similar overall food intake and activity levels. On the HF diet, CK1εtau/tau mice on a 20 hr LD cycle were protected against the development of HF diet-induced excess weight gain. These results provide additional evidence supporting a link between circadian rhythms and energy regulation at the genetic level, particularly highlighting CK1ε involved in the integration of circadian biology and metabolic physiology. PMID:27144030

  14. Placental glucose and amino acid transport in calorie-restricted wild-type and Glut3 null heterozygous mice.

    PubMed

    Ganguly, Amit; Collis, Laura; Devaskar, Sherin U

    2012-08-01

    Calorie restriction (CR) decreased placenta and fetal weights in wild-type (wt) and glucose transporter (Glut) 3 heterozygous null (glut3(+/-)) mice. Because placental nutrient transport is a primary energy determinant of placentofetal growth, we examined key transport systems. Maternal CR reduced intra- and transplacental glucose and leucine transport but enhanced system A amino acid transport in wt mice. These transport perturbations were accompanied by reduced placental Glut3 and leucine amino acid transporter (LAT) family member 2, no change in Glut1 and LAT family member 1, but increased sodium coupled neutral amino acid transporter (SNAT) and SNAT2 expression. We also noted decreased total and active phosphorylated forms of mammalian target of rapamycin, which is the intracellular nutrient sensor, the downstream total P70S6 kinase, and pS6 ribosomal protein with no change in total and phosphorylated 4E-binding protein 1. To determine the role of placental Glut3 in mediating CR-induced placental transport changes, we next investigated the effect of gestational CR in glut3(+/-) mice. In glut3(+/-) mice, a key role of placental Glut3 in mediating transplacental and intraplacental glucose transport was established. In addition, reduced Glut3 results in a compensatory increase of leucine and system A transplacental transport. On the other hand, diminished Glut3-mediated intraplacental glucose transport reduced leucine transport and mammalian target of rapamycin and preserved LAT and enhancing SNAT. CR in glut3(+/-) mice further reduced transplacental glucose transport and enhanced system A amino acid transport, although the increased leucine transport was lost. In addition, increased Glut3 was seen and preserved Glut1, LAT, and SNAT. These placental changes collectively protect survival of wt and glut3(+/-) fetuses against maternal CR-imposed reduction of macromolecular nutrients.

  15. Placental Glucose and Amino Acid Transport in Calorie-Restricted Wild-Type and Glut3 Null Heterozygous Mice

    PubMed Central

    Ganguly, Amit; Collis, Laura

    2012-01-01

    Calorie restriction (CR) decreased placenta and fetal weights in wild-type (wt) and glucose transporter (Glut) 3 heterozygous null (glut3+/−) mice. Because placental nutrient transport is a primary energy determinant of placentofetal growth, we examined key transport systems. Maternal CR reduced intra- and transplacental glucose and leucine transport but enhanced system A amino acid transport in wt mice. These transport perturbations were accompanied by reduced placental Glut3 and leucine amino acid transporter (LAT) family member 2, no change in Glut1 and LAT family member 1, but increased sodium coupled neutral amino acid transporter (SNAT) and SNAT2 expression. We also noted decreased total and active phosphorylated forms of mammalian target of rapamycin, which is the intracellular nutrient sensor, the downstream total P70S6 kinase, and pS6 ribosomal protein with no change in total and phosphorylated 4E-binding protein 1. To determine the role of placental Glut3 in mediating CR-induced placental transport changes, we next investigated the effect of gestational CR in glut3+/− mice. In glut3+/− mice, a key role of placental Glut3 in mediating transplacental and intraplacental glucose transport was established. In addition, reduced Glut3 results in a compensatory increase of leucine and system A transplacental transport. On the other hand, diminished Glut3-mediated intraplacental glucose transport reduced leucine transport and mammalian target of rapamycin and preserved LAT and enhancing SNAT. CR in glut3+/− mice further reduced transplacental glucose transport and enhanced system A amino acid transport, although the increased leucine transport was lost. In addition, increased Glut3 was seen and preserved Glut1, LAT, and SNAT. These placental changes collectively protect survival of wt and glut3+/− fetuses against maternal CR-imposed reduction of macromolecular nutrients. PMID:22700768

  16. Altered Trabecular Bone Structure and Delayed Cartilage Degeneration in the Knees of Collagen VI Null Mice

    PubMed Central

    Christensen, Susan E.; Coles, Jeffrey M.; Zelenski, Nicole A.; Furman, Bridgette D.; Leddy, Holly A.; Zauscher, Stefan; Bonaldo, Paolo; Guilak, Farshid

    2012-01-01

    Mutation or loss of collagen VI has been linked to a variety of musculoskeletal abnormalities, particularly muscular dystrophies, tissue ossification and/or fibrosis, and hip osteoarthritis. However, the role of collagen VI in bone and cartilage structure and function in the knee is unknown. In this study, we examined the role of collagen VI in the morphology and physical properties of bone and cartilage in the knee joint of Col6a1−/− mice by micro-computed tomography (microCT), histology, atomic force microscopy (AFM), and scanning microphotolysis (SCAMP). Col6a1−/− mice showed significant differences in trabecular bone structure, with lower bone volume, connectivity density, trabecular number, and trabecular thickness but higher structure model index and trabecular separation compared to Col6a1+/+ mice. Subchondral bone thickness and mineral content increased significantly with age in Col6a1+/+ mice, but not in Col6a1−/− mice. Col6a1−/− mice had lower cartilage degradation scores, but developed early, severe osteophytes compared to Col6a1+/+mice. In both groups, cartilage roughness increased with age, but neither the frictional coefficient nor compressive modulus of the cartilage changed with age or genotype, as measured by AFM. Cartilage diffusivity, measured via SCAMP, varied minimally with age or genotype. The absence of type VI collagen has profound effects on knee joint structure and morphometry, yet minimal influences on the physical properties of the cartilage. Together with previous studies showing accelerated hip osteoarthritis in Col6a1−/− mice, these findings suggest different roles for collagen VI at different sites in the body, consistent with clinical data. PMID:22448243

  17. Epac2a-null mice exhibit obesity-prone nature more susceptible to leptin resistance

    PubMed Central

    Hwang, M; Go, Y; Park, J-H; Shin, S-K; Song, S E; Oh, B-C; Im, S-S; Hwang, I; Jeon, Y H; Lee, I-K; Seino, S; Song, D-K

    2017-01-01

    Background: The exchange protein directly activated by cAMP (Epac), which is primarily involved in cAMP signaling, has been known to be essential for controlling body energy metabolism. Epac has two isoforms: Epac1 and Epac2. The function of Epac1 on obesity was unveiled using Epac1 knockout (KO) mice. However, the role of Epac2 in obesity remains unclear. Methods: To evaluate the role of Epac2 in obesity, we used Epac2a KO mice, which is dominantly expressed in neurons and endocrine tissues. Physiological factors related to obesity were analyzed: body weight, fat mass, food intake, plasma leptin and adiponectin levels, energy expenditure, glucose tolerance, and insulin and leptin resistance. To determine the mechanism of Epac2a, mice received exogenous leptin and then hypothalamic leptin signaling was analyzed. Results: Epac2a KO mice appeared to have normal glucose tolerance and insulin sensitivity until 12 weeks of age, but an early onset increase of plasma leptin levels and decrease of plasma adiponectin levels compared with wild-type mice. Acute leptin injection revealed impaired hypothalamic leptin signaling in KO mice. Consistently, KO mice fed a high-fat diet (HFD) were significantly obese, presenting greater food intake and lower energy expenditure. HFD-fed KO mice were also characterized by greater impairment of hypothalamic leptin signaling and by weaker leptin-induced decrease in food consumption compared with HFD-fed wild-type mice. In wild-type mice, acute exogenous leptin injection or chronic HFD feeding tended to induce hypothalamic Epac2a expression. Conclusions: Considering that HFD is an inducer of hypothalamic leptin resistance and that Epac2a functions in pancreatic beta cells during demands of greater work load, hypothalamic Epac2a may have a role in facilitating leptin signaling, at least in response to higher metabolic demands. Thus, our data indicate that Epac2a is critical for preventing obesity and thus Epac2a activators may be used to

  18. Delayed liver regeneration after partial hepatectomy in adipose differentiation related protein-null mice

    PubMed Central

    Kohjima, Motoyuki; Tsai, Tsung-Huang; Tackett, Bryan C.; Thevananther, Sundararajah; Li, Lan; Chang, Benny Hung-Junn; Chan, Lawrence

    2014-01-01

    Background & Aims Adult hepatocytes undergo cell cycle progression and proliferation in response to partial hepatectomy (PH). Transient lipid accumulation within hepatocytes preceding the peak proliferative phase is a characteristic feature of regenerating livers. However, the molecular mediators and mechanisms responsible for lipid accumulation in regenerating livers are not well understood. Adipose differentiation related protein (ADRP; Plin2) regulates hepatic triglyceride storage and Plin2-deficient (Plin2−/−) mice have significantly reduced triglyceride (TG) content in the liver. We sought to determine the functional significance of PLIN2 in liver regeneration in response to PH and toxic liver injury and examined whether absence of Plin2 expression modulates hepatocyte proliferation and liver regeneration. Methods We subjected wild-type (WT) and Plin2−/− mice to 70% PH or acute carbon tetrachloride (CCL4) treatment and examined the hepatic lipid content, the expression profile of lipid metabolism-related genes, the rate of cellular proliferation and the dynamics of liver regeneration in the treated animals. Results In response to PH, Plin2−/− mice showed decreased hepatic triglyceride accumulation and delayed cell cycle progression, which was associated with impaired liver regeneration. Fatty acid (FA) synthesis and lipid transfer gene expression profile were comparable between Plin2−/− and wild-type mice, while VLDL secretion rate was higher in the Plin2−/− mice. Downregulated β-oxidation and reduced cytosolic FA level in Plin2−/− mice may have contributed to the attenuation of the liver regeneration capacity in these animals. In parallel experiments, we also observed attenuated hepatic lipid accumulation and proliferation in response to CCl4-mediated acute toxic liver injury in Plin2−/− mice. Conclusions We conclude that PLIN2-mediated lipid accumulation and utilization by the liver is important for efficient liver regeneration

  19. Characterizing the role of endothelin-1 in the progression of cardiac hypertrophy in aryl hydrocarbon receptor (AhR) null mice

    SciTech Connect

    Lund, Amie K.; Goens, M. Beth; Nunez, Bethany A.; Walker, Mary K. . E-mail: mkwalker@unm.edu

    2006-04-15

    The aryl hydrocarbon receptor (AhR) is a ligand-activated transcription factor characterized to play a role in detection and adaptation to environmental stimuli. Genetic deletion of AhR results in hypertension, and cardiac hypertrophy and fibrosis, associated with elevated plasma angiotensin II (Ang II) and endothelin-1 (ET-1), thus AhR appears to contribute to cardiovascular homeostasis. In these studies, we tested the hypothesis that ET-1 mediates cardiovascular pathology in AhR null mice via ET{sub A} receptor activation. First, we determine the time courses of cardiac hypertrophy, and of plasma and tissue ET-1 expression in AhR wildtype and null mice. AhR null mice exhibited increases in heart-to-body weight ratio and age-related expression of cardiac hypertrophy markers, {beta}-myosin heavy chain ({beta}-MHC), and atrial natriuretic factor (ANF), which were significant at 2 months. Similarly, plasma and tissue ET-1 expression was significantly elevated at 2 months and increased further with age. Second, AhR null mice were treated with ET{sub A} receptor antagonist, BQ-123 (100 nmol/kg/day), for 7, 28, or 58 days and blood pressure, cardiac fibrosis, and cardiac hypertrophy assessed, respectively. BQ-123 for 7 days significantly reduced mean arterial pressure in conscious, catheterized mice. BQ-123 for 28 days significantly reduced the histological appearance of cardiac fibrosis. Treatment for 58 days significantly reduced cardiac mass, assessed by heart weight, echocardiography, and {beta}-MHC and ANF expression; and reduced cardiac fibrosis as determined by osteopontin and collagen I mRNA expression. These findings establish ET-1 and the ET{sub A} receptor as primary determinants of hypertension and cardiac pathology in AhR null mice.

  20. Ciliary neurotrophic factor is not required for terminal sprouting and compensatory reinnervation of neuromuscular synapses: Re-evaluation of CNTF null mice

    PubMed Central

    Wright, Megan C.; Son, Young-Jin

    2007-01-01

    Loss of synaptic activity or innervation induces sprouting of intact motor nerve terminals that adds or restores nerve-muscle connectivity. Ciliary neurotrophic factor (CNTF) and terminal Schwann cells (tSCs) have been implicated as molecular and cellular mediators of the compensatory process. We wondered if the previously reported lack of terminal sprouting in CNTF null mice was due to abnormal reactivity of tSCs. To this end, we examined nerve terminal and tSC responses in CNTF null mice using experimental systems that elicited extensive sprouting in wildtype mice. Contrary to the previous report, we found that motor nerve terminals in the null mice sprout extensively in response to major sprouting-stimuli such as exogenously applied CNTF per se, botulinum toxin-elicited paralysis, and partial denervation by L4 spinal root transection. In addition, the number, length and growth patterns of terminal sprouts, and the extent of reinnervation by terminal or nodal sprouts, were similar in wildtype and null mice. tSCs in the null mice were also reactive to the sprouting-stimuli, elaborating cellular processes that accompanied terminal sprouts or guided reinnervation of denervated muscle fibers. Lastly, CNTF was absent in quiescent tSCs in intact, wildtype muscles and little if any was detected in reactive tSCs in denervated muscles. Thus, CNTF is not required for induction of nerve terminal sprouting, for reactivation of tSCs, and for compensatory reinnervation after nerve injury. We interpret these results to support the notion that compensatory sprouting in adult muscles is induced primarily by contact-mediated mechanisms, rather than by diffusible factors. PMID:17445802

  1. Pancreas-specific aquaporin 12 null mice showed increased susceptibility to caerulein-induced acute pancreatitis.

    PubMed

    Ohta, Eriko; Itoh, Tomohiro; Nemoto, Tomomi; Kumagai, Jiro; Ko, Shigeru B H; Ishibashi, Kenichi; Ohno, Mayuko; Uchida, Keiko; Ohta, Akihito; Sohara, Eisei; Uchida, Shinichi; Sasaki, Sei; Rai, Tatemitsu

    2009-12-01

    Aquaporin 12 (AQP12) is the most recently identified member of the mammalian AQP family and is specifically expressed in pancreatic acinar cells. In vitro expression studies have revealed that AQP12 is localized at intracellular sites. To determine the physiological roles of AQP12 in the pancreas, we generated knockout mice for this gene (AQP12-KO). No obvious differences were observed under normal conditions between wild-type (WT) and AQP12-KO mice in terms of growth, blood chemistry, pancreatic fluid content, or histology. However, when we induced pancreatitis through the administration of a cholecystokinin-8 (CCK-8) analog, the AQP12-KO mice showed more severe pathological damage to this organ than WT mice. Furthermore, when we analyzed exocytosis in the pancreatic acini using a two-photon excitation imaging method, the results revealed larger exocytotic vesicles (vacuoles) in the acini of AQP12-KO mice at a high CCK-8 dose (100 nM). From these results, we conclude that AQP12 may function in the mechanisms that control the proper secretion of pancreatic fluid following rapid and intense stimulation.

  2. Soluble maltase-glucoamylase is alternatively spliced and secreted by Paneth and goblet cells in enterocyte maltase-glucoamylase Null mice

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Starch is the major energy source in the mouse diet. Knockout of intestinal membrane-bound maltase-glucoamylase (Mgam) in mice revealed presence of soluble starch digesting activity in Null jejunum and ileum that is secreted by Paneth and goblet cells. Hypotheses: 1. WT have two Mgam mRNAs, Mgamme...

  3. Ghrelin Receptor Deficiency does not Affect Diet-Induced Atherosclerosis in Low-Density Lipoprotein Receptor-Null Mice

    PubMed Central

    Habegger, Kirk M.; Grant, Erin; Pfluger, Paul Thomas; Perez-Tilve, Diego; Daugherty, Alan; Bruemmer, Dennis; Tschöp, Matthias H.; Hofmann, Susanna M.

    2011-01-01

    Objective: Ghrelin, a stomach-derived, secreted peptide, and its receptor (growth hormone secretagogue receptor, GHSR) are known to modulate food intake and energy homeostasis. The ghrelin system is also expressed broadly in cardiovascular tissues. Since ghrelin has been associated with anti-inflammatory and anti-atherogenic properties, but is also well known to promote obesity and impair glucose metabolism, we investigated whether ghrelin has any impact on the development of atherosclerosis. The hypothesis that endogenous ghrelin signaling may be involved in atherosclerosis has not been tested previously. Methods and Results: We crossed ghrelin receptor knockout mice (GHSr−/−) into a low-density lipoprotein receptor-null (Ldlr−/−) mouse line. In this model, atherosclerotic lesions were promoted by feeding a high-fat, high-cholesterol Western-type diet for 13 months, following a standard protocol. Body composition and glucose homeostasis were similar between Ldlr−/− and Ldlr/GHSR−/−ko mice throughout the study. Absence or presence of GHSr did not alter the apolipoprotein profile changes in response to diet exposure on an LDLRko background. Atherosclerotic plaque volume in the aortic arch and thoracic aorta were also not affected differentially in mice without ghrelin signaling due to GHSR gene disruption as compared to control LDLRko littermates. In light of the associations reported for ghrelin with cardiovascular disease in humans, the lack of a phenotype in these loss-of-function studies in mice suggests no direct role for endogenous ghrelin in either the inhibition or the promotion of diet-induced atherosclerosis. Conclusion: These data indicate that, surprisingly, the complex and multifaceted actions of endogenous ghrelin receptor mediated signaling on the cardiovascular system have minimal direct impact on atherosclerotic plaque progression as based on a loss-of-function mouse model of the disease. PMID:22649381

  4. GENE PROFILING IN WILD TYPE AND PPARÁ NULL MICE EXPOSED TO PFOA

    EPA Science Inventory

    Perflurooctanoic acid (PFOA) is a perfluoroalkyl acid used in a variety of commercial applications. Concerns have been raised because PFOA is ubiquitous in the environment and can be detected in human tissues. PFOA is a rodent carcinogen and a developmental toxicant in mice. W...

  5. Nerve injury induces robust allodynia and ectopic discharges in Nav1.3 null mutant mice

    PubMed Central

    Nassar, Mohammed A; Baker, Mark D; Levato, Alessandra; Ingram, Rachel; Mallucci, Giovanna; McMahon, Stephen B; Wood, John N

    2006-01-01

    Changes in sodium channel activity and neuronal hyperexcitability contribute to neuropathic pain, a major clinical problem. There is strong evidence that the re-expression of the embryonic voltage-gated sodium channel subunit Nav1.3 underlies neuronal hyperexcitability and neuropathic pain. Here we show that acute and inflammatory pain behaviour is unchanged in global Nav1.3 mutant mice. Surprisingly, neuropathic pain also developed normally in the Nav1.3 mutant mouse. To rule out any genetic compensation mechanisms that may have masked the phenotype, we investigated neuropathic pain in two conditional Nav1.3 mutant mouse lines. We used Nav1.8-Cre mice to delete Nav1.3 in nociceptors at E14 and NFH-Cre mice to delete Nav1.3 throughout the nervous system postnatally. Again normal levels of neuropathic pain developed after nerve injury in both lines. Furthermore, ectopic discharges from damaged nerves were unaffected by the absence of Nav1.3 in global knock-out mice. Our data demonstrate that Nav1.3 is neither necessary nor sufficient for the development of nerve-injury related pain. PMID:17052333

  6. Lipid abnormalities in alpha/beta2-syntrophin null mice are independent from ABCA1

    PubMed Central

    Hebel, Tobias; Eisinger, Kristina; Neumeier, Markus; Rein-Fischboeck, Lisa; Pohl, Rebekka; Meier, Elisabeth M.; Boettcher, Alfred; Froehner, Stanley C.; Adams, Marvin E.; Liebisch, Gerhard; Krautbauer, Sabrina; Buechler, Christa

    2015-01-01

    The syntrophins alpha (SNTA) and beta 2 (SNTB2) are molecular adaptor proteins shown to stabilize ABCA1, an essential regulator of HDL cholesterol. Furthermore, SNTB2 is involved in glucose stimulated insulin release. Hyperglycemia and dyslipidemia are characteristic features of the metabolic syndrome, a serious public health problem with rising prevalence. Therefore, it is important to understand the role of the syntrophins herein. Mice deficient for both syntrophins (SNTA/B2−/−) have normal insulin and glucose tolerance, hepatic ABCA1 protein and cholesterol. When challenged with a HFD, wild type and SNTA/B2−/− mice have similar weight gain, adiposity, serum and liver triglycerides. Hepatic ABCA1, serum insulin and insulin sensitivity are normal while glucose tolerance is impaired. Liver cholesterol is reduced, and expression of SREBP2 and HMG-CoA-R is increased in the knockout mice. Scavenger receptor-BI (SR-BI) protein is strongly diminished in the liver of SNTA/B2−/− mice while SR-BI binding protein NHERF1 is not changed and PDZK1 is even induced. Knock-down of SNTA, SNTB2 or both has no effect on hepatocyte SR-BI and PDZK1 proteins. Further, SR-BI levels are not reduced in brown adipose tissue of SNTA/B2−/− mice excluding that syntrophins directly stabilize SR-BI. SR-BI stability is regulated by MAPK and phosphorylated ERK2 is induced in the liver of the knock-out mice. Blockage of ERK activity upregulates hepatocyte SR-BI showing that increased MAPK activity contributes to low SR-BI. Sphingomyelin which is well described to regulate cholesterol metabolism is reduced in the liver and serum of the knock-out mice while the size of serum lipoproteins is not affected. Current data exclude a major function of these syntrophins in ABCA1 activity and insulin release but suggest a role in regulating glucose uptake, ERK and SR-BI levels, and sphingomyelin metabolism in obesity. PMID:25625330

  7. PKCβ Promotes Vascular inflammation and Acceleration of Atherosclerosis in Diabetic ApoE Null Mice

    PubMed Central

    Kong, Linghua; Shen, Xiaoping; Lin, Lili; Leitges, Michael; Rosario, Rosa; Zou, Yu Shan; Yan, Shi Fang

    2013-01-01

    Objective Diabetic subjects are at high risk for developing atherosclerosis through a variety of mechanisms. As the metabolism of glucose results in production of activators of protein kinase C (PKC)β, it was logical to investigate the role of PKCβ in modulation of atherosclerosis in diabetes. Approach and Results ApoE−/− and PKCβ −/−/ApoE−/− mice were rendered diabetic with streptozotocin. Quantification of atherosclerosis, gene expression profiling or analysis of signaling molecules was performed on aortic sinus or aortas from diabetic mice. Diabetes-accelerated atherosclerosis increased the level of phosphorylated ERK1/2 and JNK mitogen activated protein (MAP) kinases and augmented vascular expression of inflammatory mediators, as well as increased monocyte/macrophage infiltration and CD11c+ cells accumulation in diabetic ApoE−/− mice; processes which were diminished in diabetic PKCβ −/−/ApoE−/− mice. In addition, pharmacological inhibition of PKCβ reduced atherosclerotic lesion size in diabetic ApoE−/− mice. In vitro, the inhibitors of PKCβ and ERK1/2, as well as small interfering RNA (siRNA) to Egr-1 significantly decreased high glucose-induced expression of CD11c (Itgax), chemokine (C-C motif) ligand 2 (CCL2) and interleukin (IL)-1β in U937 macrophages. Conclusions These data link enhanced activation of PKCβ to accelerated diabetic atherosclerosis via a mechanism that includes modulation of gene transcription and signal transduction in the vascular wall; processes that contribute to acceleration of vascular inflammation and atherosclerosis in diabetes. Our results uncover a novel role for PKCβ in modulating CD11c expression and inflammatory response of macrophages in the development of diabetic atherosclerosis. These findings support PKCβ activation as a potential therapeutic target for prevention and treatment of diabetic atherosclerosis. PMID:23766264

  8. Comparative metabolism of methacrylonitrile and acrylonitrile to cyanide using cytochrome P4502E1 and microsomal epoxide hydrolase-null mice

    SciTech Connect

    El Hadri, L.; Chanas, B.; Ghanayem, B.I. . E-mail: ghanayem@niehs.nih.gov

    2005-06-01

    Methacrylonitrile (MAN) and acrylonitrile (AN) are metabolized via glutathione (GSH) conjugation or epoxide formation. We have recently shown that CYP2E1 is essential for AN epoxidation and subsequent cyanide liberation. Current studies were designed to compare the enzymatic basis of MAN vs. AN metabolism to cyanide using wild-type (WT), CYP2E1-, and mEH-null mice. Mice received a single gavage dose of 0.047, 0.095, 0.19, or 0.38 mmol/kg of MAN or AN, and blood cyanide was measured at 1 or 3 h later. Blood cyanide levels in WT mice treated with AN or MAN were dose and time dependent. At equimolar doses, significantly higher levels of cyanide were detected in the blood of MAN- vs. AN-treated mice. Further, while significant reduction in blood cyanide levels occurred in MAN-treated CYP2E1-null vs. WT mice, AN metabolism to cyanide was largely abolished in CYP2E1-null mice. Pretreatment of mice with 1-aminobenzotriazole (ABT, CYP inhibitor) demonstrated that CYPs other than CYP2E1 also contribute to MAN metabolism to cyanide. Blood cyanide levels in mEH-null mice treated with aliphatic nitriles are generally lower than levels in similarly treated WT mice. Western blot analysis showed that expression of sEH was greater in male vs. female mice. The role of various epoxide hydrolases (EHs) in the production of cyanide from aliphatic nitriles is apparently structure and dose dependent. Regardless of genotype, significantly higher levels of cyanide were measured in the blood of male vs. female mice treated with MAN or AN. In conclusion, these data showed that (1) at equimolar doses, higher blood cyanide levels were detected in mice treated with MAN vs. AN; (2) while CYP2E1 is the only enzyme responsible for AN metabolism to cyanide, other CYPs also contribute to MAN metabolism; and (3) significantly higher levels of cyanide were measured in the blood of male vs. female treated with either nitrile. Higher blood cyanide levels in male vs. female mice and in MAN- vs. AN

  9. A role for TGFbeta1 in langerhans cell biology. Further characterization of the epidermal Langerhans cell defect in TGFbeta1 null mice.

    PubMed Central

    Borkowski, T A; Letterio, J J; Mackall, C L; Saitoh, A; Wang, X J; Roop, D R; Gress, R E; Udey, M C

    1997-01-01

    Previous studies of TGFbeta1 null (-/-) mice indicated that the epidermis was devoid of Langerhans cells (LC) and that the LC deficiency was not secondary to the inflammation that is the dominant feature of the -/- phenotype (Borkowski, T.A., J.J. Letterio, A.G. Farr, and M.C. Udey. 1996. J. Exp. Med. 184:2417-2422). Herein, we demonstrate that dendritic cells could be expanded from the bone marrow of -/- mice and littermate controls. Bone marrow from -/- mice also gave rise to LC after transfer into lethally irradiated recipients. Thus, the LC defect in TGFbeta1 null mice does not result from an absolute deficiency in bone marrow precursors, and paracrine TGFbeta1 production is sufficient for LC development. Several approaches were used to assess the suitability of -/- skin for LC localization. A survey revealed that although a number of cytokine mRNAs were expressed de novo, mRNAs encoding proinflammatory cytokines known to mobilize LC from epidermis (IL-1 and TNFalpha) were not strikingly overrepresented in -/- skin. In addition, bone marrow-derived LC populated full-thickness TGFbeta1 null skin after engraftment onto BALB/c nu/nu recipients. Finally, the skin of transgenic mice expressing a truncated loricrin promoter-driven dominant-negative TGFbeta type II receptor contained normal numbers of LC. Because TGFbeta1 signaling in these mice is disrupted only in keratinocytes and the keratinocyte hyperproliferative component of the TGFbeta1 -/- phenotype is reproduced, these results strongly suggest that the LC defect in TGFbeta1 null mice is not due to an epidermal abnormality but reflects a requirement of murine LC (or their precursors) for TGFbeta1. PMID:9239404

  10. Glutathione deficiency in Gclm null mice results in complex I inhibition and dopamine depletion following paraquat administration.

    PubMed

    Liang, Li-Ping; Kavanagh, Terrance J; Patel, Manisha

    2013-08-01

    Depletion of glutathione has been shown to occur in autopsied brains of patients with Parkinson's disease (PD) and in animal models of PD. The goal of this study was to determine whether chronic glutathione (GSH) deficiency per se resulted in complex I inhibition and/or dopamine depletion and whether these indices were further potentiated by aging or administration of paraquat, a redox-cycling herbicide that produces a PD-like neurodegeneration model in rodents (Brooks, A. I., Chadwick, C. A., Gelbard, H. A., Cory-Slechta, D. A., and Federoff, H. J. [1999]. Paraquat elicited neurobehavioral syndrome caused by dopaminergic neuron loss. Brain Res. 823, 1-10; McCormack, A. L., Thiruchelvam, M., Manning-Bog, A. B., Thiffault, C., Langston, J. W., Cory-Slechta, D. A., and Di Monte, D. A. [2002]. Environmental risk factors and Parkinson's disease: Selective degeneration of nigral dopaminergic neurons caused by the herbicide paraquat. Neurobiol. Dis. 10, 119-127.) Deletion of the rate-limiting GSH synthesis gene, glutamate-cysteine ligase modifier subunit (Gclm), leads to significantly lower GSH concentrations in all tissues including brain. Gclm null (Gclm (-/-)) mice provide a model of prolonged GSH depletion to explore the relationship between GSH, complex I inhibition, and dopamine loss in vivo. Despite ~60% depletion of brain GSH in Gclm (-/-) mice of ages 3-5 or 14-16 months, striatal complex I activity, dopamine levels, 3-nitrotyroine/tyrosine ratios, aconitase activity, and CoASH remained unchanged. Administration of paraquat (10mg/kg, twice/week, 3 weeks) to 3- to 5-month-old Gclm (-/-) mice resulted in significantly decreased aconitase activity, complex I activity, and dopamine levels but not in 3- to 5-month-old Gclm (+/+) mice. Furthermore, paraquat-induced inhibition of complex I and aconitase activities in Gclm (-/-) mice was observed in the striatum but not in the cortex. The results suggest that chronic deficiency of GSH in Gclm (-/-) mice was not

  11. Genetically null mice reveal a central role for epidermal growth factor receptor in the differentiation of the hair follicle and normal hair development.

    PubMed Central

    Hansen, L. A.; Alexander, N.; Hogan, M. E.; Sundberg, J. P.; Dlugosz, A.; Threadgill, D. W.; Magnuson, T.; Yuspa, S. H.

    1997-01-01

    Mice harboring a targeted disruption of the epidermal growth factor receptor (EGFR) allele exhibit a severely disorganized hair follicle phenotype, fuzzy coat, and systemic disease resulting in death before 3 weeks. This skin phenotype was reproduced in whole skin grafts and in grafts of EGFR null hair follicle buds onto nude mice, providing a model to evaluate the natural evolution of skin lacking the EGFR. Hair follicles in grafts of null skin did not progress from anagen to telogen and scanning electron micrografts revealed wavy, flattened hair fibers with cuticular abnormalities. Many of the EGFR null hair follicles in the grafted skin were consumed by an inflammatory reaction resulting in complete hair loss in 67% of the grafts by 10 weeks. Localization of follicular differentiation markers including keratin 6, transglutaminase, and the hair keratins mHa2 and hacl-1 revealed a pattern of premature differentiation within the null hair follicles. In intact EGFR null mice, proliferation in the interfollicular epidermis, but not hair follicles, was greatly decreased in the absence of EGFR. In contrast, grafting of EGFR null skin resulted in a hyperplastic response in the epidermis that did not resolve even after 10 weeks, although the wound-induced hyperplasia in EGFR wild-type grafts had resolved within 3 to 4 weeks. Thus, epithelial expression of the EGFR has complex functions in the skin. It is important in delaying follicular differentiation, may serve to protect the hair follicle from immunological reactions, and modifies both normal and wound-induced epidermal proliferation but seems dispensable for follicular proliferation. Images Figure 1 Figure 2 Figure 3 Figure 4 Figure 5 Figure 6 Figure 7 Figure 8 PMID:9176390

  12. Aged PrP null mice show defective processing of neuregulins in the peripheral nervous system.

    PubMed

    Benvegnù, Stefano; Gasperini, Lisa; Legname, Giuseppe

    2011-05-01

    A prion, a protease-resistant conformer of the cellular prion protein (PrP(C)), is the causative agent of transmissible spongiform encephalopathies or prion diseases. While this property is well established for the aberrantly folded protein, the physiological function of PrP(C) remains elusive. Among different putative functions, the non-pathogenic protein isoform PrP(C) is involved in several cellular processes. Here, we show that PrP(C) regulates the cleavage of neuregulin-1 proteins (NRG1). Neuregulins provide key axonal signals that regulate several processes, including glial cells proliferation, survival and myelination. Interestingly, mice devoid of PrP(C) (Prnp⁰/⁰) were recently shown to have a late-onset demyelinating disease in the peripheral nervous system (PNS) but not in the central nervous system (CNS). We found that NRG1 processing is developmentally regulated in the PNS and, by comparing wildtype and Prnp⁰/⁰ mice, that PrP(C) influences NRG1 processing in old, but not in young, animals. In addition, we found that also the processing of neuregulin-3, another neuregulin family member, is altered in the PNS of Prnp⁰/⁰ mice. These differences in neuregulin proteins processing are not paralleled in the CNS, thus suggesting a different cellular function for PrP(C) between the CNS and the PNS.

  13. Lacrimal gland inflammation is responsible for ocular pathology in TGF-beta 1 null mice.

    PubMed Central

    McCartney-Francis, N. L.; Mizel, D. E.; Frazier-Jessen, M.; Kulkarni, A. B.; McCarthy, J. B.; Wahl, S. M.

    1997-01-01

    Mice homozygous for a nonfunctional transforming growth factor-beta 1 gene develop rampant inflammation in vital organs that contributes to a shortened life span. The presence of circulating anti-nuclear anti-bodies, immune deposits in tissues, leukocyte infiltration, and increased major histocompatibility complex antigen expression resembles an autoimmune-like syndrome. One of the overt symptoms that appears in these mice lacking transforming growth factor-beta 1 is the development of dry crusty eyes that close persistently as their health declines. Histologically, the eyes appear normal with little or no inflammation. However, inflammatory lesions, predominantly lymphocytic, develop in the lacrimal glands, disrupting their structure and function and severely limiting their ability to generate tears. This histopathology and aberrant function mimic that of Sjögren's syndrome, a human autoimmune disease characterized by dry eyes and dry mouth. Impeding the leukocyte infiltration into the glands with synthetic fibronectin peptides, which block adhesion, not only prevents the inflammatory pathology but also prevents the persistent eye closure characteristic of these mice. Images Figure 1 Figure 2 Figure 3 Figure 4 Figure 5 PMID:9358754

  14. Mice with an NaV1.4 sodium channel null allele have latent myasthenia, without susceptibility to periodic paralysis.

    PubMed

    Wu, Fenfen; Mi, Wentao; Fu, Yu; Struyk, Arie; Cannon, Stephen C

    2016-06-01

    Over 60 mutations of SCN4A encoding the NaV1.4 sodium channel of skeletal muscle have been identified in patients with myotonia, periodic paralysis, myasthenia, or congenital myopathy. Most mutations are missense with gain-of-function defects that cause susceptibility to myotonia or periodic paralysis. Loss-of-function from enhanced inactivation or null alleles is rare and has been associated with myasthenia and congenital myopathy, while a mix of loss and gain of function changes has an uncertain relation to hypokalaemic periodic paralysis. To better define the functional consequences for a loss-of-function, we generated NaV1.4 null mice by deletion of exon 12. Heterozygous null mice have latent myasthenia and a right shift of the force-stimulus relation, without evidence of periodic paralysis. Sodium current density was half that of wild-type muscle and no compensation by retained expression of the foetal NaV1.5 isoform was detected. Mice null for NaV1.4 did not survive beyond the second postnatal day. This mouse model shows remarkable preservation of muscle function and viability for haploinsufficiency of NaV1.4, as has been reported in humans, with a propensity for pseudo-myasthenia caused by a marginal Na(+) current density to support sustained high-frequency action potentials in muscle.

  15. Passive Viscoelastic Properties of Costameres in EDL Skeletal Muscle in Normal and Dystrophin Null Mice

    NASA Astrophysics Data System (ADS)

    García-Pelagio, Karla P.; Bloch, Robert J.; Ortega, Alicia; Gonzalez-Serratos, Hugo

    2008-08-01

    Costameres at the sarcolemmal skeletal myofibers transmit the lateral force generated by myofibrils from them to the extracellular matrix. We used an elastimeter method by which sucking pressure is applied through a micropipette to the surface membrane of single mice myofibers of the Extensor digitorum longus to measure the viscoelasticity of the sarcolemma-costamere complex as a function of sarcomere length (SL). Constant suction pressure applied to the sarcolemma generated a sarcolemmal-costamere-myofibril bleb of variable height depending on the sucking pressure and SL. It took some time for the bleb to reach a stable height after applying the pressure. This time delay indicates that the sarcolemma-costamere-myofibril system acts as a viscoelastic system. We undertook the present experiments to measure the height stabilization time of the bleb at different sarcomere lengths from which we estimated the viscoelastic parameters of the system. The time course of the bleb formation was biphasic and reached a plateau between 3.5 to 1.3 and 4.9 to 3.5 min for normal and dystrophic mice respectively depending on SL of 3.0 to 5.6 μm. Based on a Maxwell-Voigt system we found the viscoelastic parameters such as viscosity, friction coefficient and the costameres (k) and sarcolemma (k1) elasticity constants.

  16. Remodelling of motor nerve terminals in demyelinating axons of periaxin null mutant mice

    PubMed Central

    Court, Felipe A; Brophy, Peter J; Ribchester, Richard R

    2015-01-01

    Myelin formation around axons increases nerve conduction velocity and regulates phenotypic characteristics of the myelinated axon. In the peripheral nervous system, demyelinating forms of hereditary Charcot-Marie-Tooth (CMT) diseases, due to Schwann-cell intrinsic molecular defects, leads to reduced nerve conduction velocity and changes in the axonal phenotype. Several mouse models of CMT diseases have been generated, allowing the study of consequences of demyelination in peripheral nerve fibres. Nevertheless, the effect of demyelination at the level of the neuromuscular synapse has been largely overlooked. Here we show that in the periaxin knock-out mice, a model of CMT condition, neuromuscular junctions develop profound morphological changes in pre-terminal region of motoraxons. These changes include extensive preterminal branches which originate in demyelinated regions of the nerve fibre and axonal swellings associated with residually-myelinated regions of the fibre. Using intracellular recording from muscle fibres we detected asynchronous failure of action potential transmission at high but not low stimulation frequencies, a phenomenon consistent with branch point failure. Taken together, our morphological and electrophysiological findings suggest that preterminal branching due to segmental demyelination near the neuromuscular synapse in periaxin KO mice may underlie phenotypic disabilities present in this mouse model of CMT disease. These results opens a new avenue of research in order to understand the cellular changes responsible for clinical disabilities in demyelinating conditions. PMID:18205176

  17. ACE2: Angiotensin II/Angiotensin-(1-7) balance in cardiorenal injury

    PubMed Central

    Varagic, Jasmina; Ahmad, Sarfaraz; Nagata, Sayaka; Ferrario, Carlos M.

    2014-01-01

    Our current recognition of the renin-angiotensin system is more convoluted than originally thought due to the discovery of multiple novel enzymes, peptides, and receptors inherent to this interactive biochemical cascade. Over the last decade angiotensin converting enzyme 2 (ACE2) has emerged as a key player in the pathophysiology of hypertension and cardiovascular and renal disease due to its pivotal role in metabolizing vasoconstrictive/hypertrophic/proliferative angiotensin II into favorable angiotensin-(1-7). This review addresses a considerable advancement in research on the role of tissue ACE2 in development and progression of hypertension and cardiorenal injury. We also summarize the results from recent clinical and experimental studies suggesting that serum or urine soluble ACE2 may serve as a novel biomarker or independent risk factor relevant for diagnosis and prognosis of cardiorenal disease. Recent proceedings on novel therapeutic approaches to enhance ACE2/angiotensin-(1-7) axis are also reviewed. PMID:24510672

  18. Dissociation of Hyperglycemia from Altered Vascular Contraction and Relaxation Mechanisms in Caveolin-1 Null Mice

    PubMed Central

    Pojoga, Luminita H.; Yao, Tham M.; Opsasnick, Lauren A.; Garza, Amanda E.; Reslan, Ossama M.; Adler, Gail K.; Williams, Gordon H.

    2014-01-01

    Hyperglycemia and endothelial dysfunction are associated with hypertension, but the specific causality and genetic underpinning are unclear. Caveolin-1 (cav-1) is a plasmalemmal anchoring protein and modulator of vascular function and glucose homeostasis. Cav-1 gene variants are associated with reduced insulin sensitivity in hypertensive individuals, and cav-1−/− mice show endothelial dysfunction, hyperglycemia, and increased blood pressure (BP). On the other hand, insulin-sensitizing therapy with metformin may inadequately control hyperglycemia while affecting the vascular outcome in certain patients with diabetes. To test whether the pressor and vascular changes in cav-1 deficiency states are related to hyperglycemia and to assess the vascular mechanisms of metformin under these conditions, wild-type (WT) and cav-1−/− mice were treated with either placebo or metformin (400 mg/kg daily for 21 days). BP and fasting blood glucose were in cav-1−/− > WT and did not change with metformin. Phenylephrine (Phe)- and KCl-induced aortic contraction was in cav-1−/− < WT; endothelium removal, the nitric-oxide synthase (NOS) blocker l-NAME (Nω-nitro-l-arginine methyl ester), or soluble guanylate cyclase (sGC) inhibitor 1H-[1,2,4]oxadiazolo[4,3-a]quinoxalin-1-one (ODQ) enhanced Phe contraction, and metformin blunted this effect. Acetylcholine-induced relaxation was in cav-1−/− > WT, abolished by endothelium removal, l-NAME or ODQ, and reduced with metformin. Nitric oxide donor sodium nitroprusside was more potent in inducing relaxation in cav-1−/− than in WT, and metformin reversed this effect. Aortic eNOS, AMPK, and sGC were in cav-1−/− > WT, and metformin decreased total and phosphorylated eNOS and AMPK in cav-1−/−. Thus, metformin inhibits both vascular contraction and NO-cGMP-dependent relaxation but does not affect BP or blood glucose in cav-1−/− mice, suggesting dissociation of hyperglycemia from altered vascular function in cav-1

  19. Oxazolone and ethanol induce colitis in non-obese diabetic-severe combined immunodeficiency interleukin-2Rγnull mice engrafted with human peripheral blood mononuclear cells

    PubMed Central

    Nolte, T; Zadeh-Khorasani, M; Safarov, O; Rueff, F; Gülberg, V; Herbach, N; Wollenberg, A; Mueller, T; Siebeck, M; Wolf, E; Gropp, R

    2013-01-01

    Oxazolone-induced colitis in mice has become a recognized model to study the efficacy of therapeutics targeting the immunological response underlying the development of inflammatory bowel disease. However, this model cannot be used when therapeutics designed to address human targets do not interact with the respective murine counterpart. In this study, we examined the induction of oxazolone mediated colitis in non-obese diabetic-severe combined immunodeficiency interleukin-2Rγnull (NOD-SCID IL2Rγnull) mice engrafted with human peripheral blood mononuclear cells (hPBMC) derived from patients suffering from ulcerative colitis (UC), atopic dermatitis (AD) and healthy volunteers. NOD-SCID IL2Rγ null mice were engrafted with hPBMC followed by challenge with oxazolone or ethanol vehicle. Mice developed the same symptoms as observed previously in immunocompetent mice. The clinical activity score increased and the colon architecture was characterized by the development of oedema, fibrosis, crypt loss and dense infiltration of predominantly T cells into the lamina propria. Fluorescence activated cell sorter (FACS) analysis of lymphocytes in the colon identified natural killer (NK) T cells as a major constituent. In contrast to studies with immunocompetent mice, we observed the same phenotype in the group challenged with ethanol vehicle. The phenotype was most pronounced in mice engrafted with PBMC derived from a patient suffering from UC, suggesting that the immunological history of the donors predisposes the engrafted mice to react to ethanol. The model described here has the potential to study the efficacy of therapeutics targeting human lymphocytes in a model which is more reflective of the human disease. In addition, it might be developed to elucidate molecular mechanisms underlying the disease. PMID:23574330

  20. Oxazolone and ethanol induce colitis in non-obese diabetic-severe combined immunodeficiency interleukin-2Rγ(null) mice engrafted with human peripheral blood mononuclear cells.

    PubMed

    Nolte, T; Zadeh-Khorasani, M; Safarov, O; Rueff, F; Gülberg, V; Herbach, N; Wollenberg, A; Mueller, T; Siebeck, M; Wolf, E; Gropp, R

    2013-05-01

    Oxazolone-induced colitis in mice has become a recognized model to study the efficacy of therapeutics targeting the immunological response underlying the development of inflammatory bowel disease. However, this model cannot be used when therapeutics designed to address human targets do not interact with the respective murine counterpart. In this study, we examined the induction of oxazolone mediated colitis in non-obese diabetic-severe combined immunodeficiency interleukin-2Rγ(null) (NOD-SCID IL2Rγ(null)) mice engrafted with human peripheral blood mononuclear cells (hPBMC) derived from patients suffering from ulcerative colitis (UC), atopic dermatitis (AD) and healthy volunteers. NOD-SCID IL2Rγ (null) mice were engrafted with hPBMC followed by challenge with oxazolone or ethanol vehicle. Mice developed the same symptoms as observed previously in immunocompetent mice. The clinical activity score increased and the colon architecture was characterized by the development of oedema, fibrosis, crypt loss and dense infiltration of predominantly T cells into the lamina propria. Fluorescence activated cell sorter (FACS) analysis of lymphocytes in the colon identified natural killer (NK) T cells as a major constituent. In contrast to studies with immunocompetent mice, we observed the same phenotype in the group challenged with ethanol vehicle. The phenotype was most pronounced in mice engrafted with PBMC derived from a patient suffering from UC, suggesting that the immunological history of the donors predisposes the engrafted mice to react to ethanol. The model described here has the potential to study the efficacy of therapeutics targeting human lymphocytes in a model which is more reflective of the human disease. In addition, it might be developed to elucidate molecular mechanisms underlying the disease.

  1. A human FSHB transgene encoding the double N-glycosylation mutant (Asn(7Δ) Asn(24Δ)) FSHβ subunit fails to rescue Fshb null mice.

    PubMed

    Wang, Huizhen; Butnev, Vladimir; Bousfield, George R; Kumar, T Rajendra

    2016-05-05

    Follicle-stimulating hormone (FSH) is a gonadotrope-derived heterodimeric glycoprotein. Both the common α- and hormone-specific β subunits contain Asn-linked N-glycan chains. Recently, macroheterogeneous FSH glycoforms consisting of β-subunits that differ in N-glycan number were identified in pituitaries of several species and subsequently the recombinant human FSH glycoforms biochemically characterized. Although chemical modification and in vitro site-directed mutagenesis studies defined the roles of N-glycans on gonadotropin subunits, in vivo functional analyses in a whole-animal setting are lacking. Here, we have generated transgenic mice with gonadotrope-specific expression of either an HFSHB(WT) transgene that encodes human FSHβ WT subunit or an HFSHB(dgc) transgene that encodes a human FSHβ(Asn7Δ 24Δ) double N-glycosylation site mutant subunit, and separately introduced these transgenes onto Fshb null background using a genetic rescue strategy. We demonstrate that the human FSHβ(Asn7Δ 24Δ) double N-glycosylation site mutant subunit, unlike human FSHβ WT subunit, inefficiently combines with the mouse α-subunit in pituitaries of Fshb null mice. FSH dimer containing this mutant FSHβ subunit is inefficiently secreted with very low levels detectable in serum. Fshb null male mice expressing HFSHB(dgc) transgene are fertile and exhibit testis tubule size and sperm number similar to those of Fshb null mice. Fshb null female mice expressing the mutant, but not WT human FSHβ subunit-containing FSH dimer are infertile, demonstrate no evidence of estrus cycles, and many of the FSH-responsive genes remain suppressed in their ovaries. Thus, HFSHB(dgc) unlike HFSHB(WT) transgene does not rescue Fshb null mice. Our genetic approach provides direct in vivo evidence that N-linked glycans on FSHβ subunit are essential for its efficient assembly with the α-subunit to form FSH heterodimer in pituitary. Our studies also reveal that N-glycans on FSHβ subunit are

  2. Decreased expression of tumor necrosis factor-alpha-stimulated gene 6 in cumulus cells of the cyclooxygenase-2 and EP2 null mice.

    PubMed

    Ochsner, Scott A; Russell, Darryl L; Day, Anthony J; Breyer, Richard M; Richards, Joanne S

    2003-03-01

    Ovulation, the release of fertilizable oocytes from mature follicles, involves tissue remodeling and increased prostaglandin (PG) signaling. Cyclooxygenase (COX)-2 is the rate-limiting enzyme during PG synthesis. Female mice null for either COX-2 or the PGE(2) receptor EP2 are infertile, show decreased ovulation, and exhibit abnormal cumulus expansion. Cumulus expansion is the production of a complex extracellular matrix surrounding the cumulus-oocyte complex (COC). Matrix components consist of hyaluronan, proteoglycans, and proteins with hyaluronan binding domains. One such hyaluronan binding protein is TNFalpha-stimulated gene 6 (TSG-6). By various methods, we show induction of TSG-6 and hyaluronan synthase-2 mRNA in ovaries of mice treated with pregnant mare serum gonadotropin and human chorionic gonadotropin. By in situ hybridization, we show that both genes are expressed in periantral mural granulosa cells and cumulus cells of the mouse ovary. Notably, RT-PCR and in situ hybridization show that TSG-6 mRNA but not hyaluronan synthase-2 mRNA expression is selectively reduced in cumulus cells of COX-2 and EP2 null mice. Western analysis further confirms that TSG-6 protein is reduced in isolated COCs but remains covalently associated with inter alpha-trypsin inhibitor in COX-2 null mice. These observations identify TSG-6 as a target of PG action and show that its production in ovulatory follicles is associated with proper formation of the cumulus-derived extracellular matrix.

  3. Neurobehavioral changes and alteration of gene expression in the brains of metallothionein-I/II null mice exposed to low levels of mercury vapor during postnatal development.

    PubMed

    Yoshida, Minoru; Honda, Masako; Watanabe, Chiho; Satoh, Masahiko; Yasutake, Akira

    2011-10-01

    This study examined the neurobehavioral changes and alteration in gene expression in the brains of metallothionein (MT)-I/II null mice exposed to low-levels of mercury vapor (Hg(0)) during postnatal development. MT-I/II null and wild-type mice were repeatedly exposed to Hg(0) at 0.030 mg/m(3) (range: 0.023-0.043 mg/m(3)), which was similar to the current threshold value (TLV), for 6 hr per day until the 20th day postpartum. The behavioral effects were evaluated with locomotor activity in the open field (OPF), learning ability in the passive avoidance response (PA) and spatial learning ability in the Morris water maze (MM) at 12 weeks of age. Hg(0)-exposed MT-I/II null mice showed a significant decrease in total locomotor activity in females, though learning ability and spatial learning ability were not affected. Immediately after Hg(0) exposure, mercury concentrations in the brain did not exceed 0.5 µg/g in any animals. Hg(0) exposure resulted in significant alterations in gene expression in the brains of both strains using DNA microarray analysis. The number of altered genes in MT-I/II null mice was higher than that in wild-type mice and calcium-calmodulin kinase II (Camk2a) involved in learning and memory in down-regulated genes was detected. These results provide useful information to elucidate the development of behavioral toxicity following low-level exposure to Hg(0).

  4. Macrophage-specific expression of human apolipoprotein E reduces atherosclerosis in hypercholesterolemic apolipoprotein E-null mice.

    PubMed Central

    Bellosta, S; Mahley, R W; Sanan, D A; Murata, J; Newland, D L; Taylor, J M; Pitas, R E

    1995-01-01

    apoE deficiency causes hyperlipidemia and premature atherosclerosis. To determine if macrophage-specific expression of apoE would decrease the extent of atherosclerosis, we expressed human apoE in macrophages of apoE-null mice (apoE-/-) and assessed the effect on lipid accumulation in cells of the arterial wall. Macrophage-specific expression of human apoE in normal mice was obtained by use of the visna virus LTR. These animals were bred with apoE-/- mice to produce animals hemizygous for expression of human apoE in macrophages in the absence of murine apoE (apoE-/-,hTgE+/0). Low levels of human apoE mRNA were present in liver and spleen and high levels in lung and peritoneal macrophages. Human apoE was secreted by peritoneal macrophages and was detected in Kupffer cells of the liver. Human apoE in the plasma of apoE-/-,hTgE+/0 mice (n = 30) was inversely correlated (P < 0.005) with the plasma cholesterol concentration. After 15 wk on a normal chow diet, atherosclerosis was assessed in apoE-/-,hTgE+/0 animals and in apoE-/-,hTgE0/0 littermates matched for plasma cholesterol level (approximately 450 mg/dl) and lipoprotein profile. There was significantly less atherosclerosis in both the aortic sinus and in the proximal aorta (P < 0.0001) in the animals expressing the human apoE transgene. In apo-E-/-,hTgE+/0 animals, which had detectable atherosclerotic lesions, human apoE was detected in the secretory apparatus of macrophage-derived foam cells in the arterial wall. The data demonstrate that expression of apoE by macrophages is antiatherogenic even in the presence of high levels of atherogenic lipoproteins. The data suggest that apoE prevents atherosclerosis by promoting cholesterol efflux from cells of the arterial wall. Images PMID:7593602

  5. Drosophila atonal fully rescues the phenotype of Math1 null mice: new functions evolve in new cellular contexts

    NASA Technical Reports Server (NTRS)

    Wang, Vincent Y.; Hassan, Bassem A.; Bellen, Hugo J.; Zoghbi, Huda Y.

    2002-01-01

    Many genes share sequence similarity between species, but their properties often change significantly during evolution. For example, the Drosophila genes engrailed and orthodenticle and the onychophoran gene Ultrabithorax only partially substitute for their mouse or Drosophila homologs. We have been analyzing the relationship between atonal (ato) in the fruit fly and its mouse homolog, Math1. In flies, ato acts as a proneural gene that governs the development of chordotonal organs (CHOs), which serve as stretch receptors in the body wall and joints and as auditory organs in the antennae. In the fly CNS, ato is important not for specification but for axonal arborization. Math1, in contrast, is required for the specification of cells in both the CNS and the PNS. Furthermore, Math1 serves a role in the development of secretory lineage cells in the gut, a function that does not parallel any known to be served by ato. We wondered whether ato and Math1 might be more functionally homologous than they appear, so we expressed Math1 in ato mutant flies and ato in Math1 null mice. To our surprise, the two proteins are functionally interchangeable.

  6. Enrichment of hematopoietic stem cells with SLAM and LSK markers for the detection of hematopoietic stem cell function in normal and Trp53 null mice

    PubMed Central

    Chen, Jichun; Ellison, Felicia M.; Keyvanfar, Keyvan; Omokaro, Stephanie O.; Desierto, Marie J.; Eckhaus, Michael A.; Young, Neal S.

    2008-01-01

    Objective To test function of hematopoietic stem cells (HSCs) in vivo in C57BL/6 (B6) and Trp53-deficient (Trp53 null) mice by using two HSC enrichment schemes. Methods Bone marrow (BM) Lin-CD41-CD48-CD150+ (SLAM, signaling lymphocyte activation molecules), Lin-CD41-CD48-CD150- (SLAM-) and Lin-Sca1+CD117+ (LSK) cells were defined by fluorescence activated cell staining (FACS). Cellular reactive oxygen species (ROS) level was also analyzed by FACS. Sorted SLAM, SLAM- and LSK cells were tested in vivo in the competitive repopulation (CR) and serial transplantation assays. Results The SLAM cell fraction was 0.0078 ± 0.0010% and 0.0135 ± 0.0010% of total BM cells in B6 and Trp53 null mice, and was highly correlated (R2 = 0.7116) with LSK cells. CD150+ BM cells also contained more ROSlow cells than did CD150- cells. B6 SLAM cells repopulated recipients much better than B6 SLAM- cells, showing high HSC enrichment. B6 SLAM cells also engrafted recipients better than Trp53 null SLAM cells in the CR and the follow-up serial transplantation assays. Similarly, LSK cells from B6 donors also had higher repopulating ability than those from Trp53 null donors. However, whole BM cells from the same B6 and Trp53 null donors showed the opposite functional trend in recipient engraftment. Conclusion Both SLAM and LSK marker sets can enrich HSCs from B6 and Trp53 mice. Deficiency of Trp53 up-regulates HSC self-renewal but causes no gain of HSC function. PMID:18562080

  7. Advancing age increases sperm chromatin damage and impairs fertility in peroxiredoxin 6 null mice

    PubMed Central

    Ozkosem, Burak; Feinstein, Sheldon I.; Fisher, Aron B.; O’Flaherty, Cristian

    2015-01-01

    Due to socioeconomic factors, more couples are choosing to delay conception than ever. Increasing average maternal and paternal age in developed countries over the past 40 years has raised the question of how aging affects reproductive success of males and females. Since oxidative stress in the male reproductive tract increases with age, we investigated the impact of advanced paternal age on the integrity of sperm nucleus and reproductive success of males by using a Prdx6−/− mouse model. We compared sperm motility, cytoplasmic droplet retention sperm chromatin quality and reproductive outcomes of young (2-month-old), adult (8-month-old), and old (20-month-old) Prdx6−/− males with their age-matched wild type (WT) controls. Absence of PRDX6 caused age-dependent impairment of sperm motility and sperm maturation and increased sperm DNA fragmentation and oxidation as well as decreased sperm DNA compaction and protamination. Litter size, total number of litters and total number of pups per male were significantly lower in Prdx6−/− males compared to WT controls. These abnormal reproductive outcomes were severely affected by age in Prdx6−/− males. In conclusion, the advanced paternal age affects sperm chromatin integrity and fertility more severely in the absence of PRDX6, suggesting a protective role of PRDX6 in age-associated decline in the sperm quality and fertility in mice. PMID:25796034

  8. Partial Restoration of CFTR Function in cftr-Null Mice following Targeted Cell Replacement Therapy.

    PubMed

    Duchesneau, Pascal; Besla, Rickvinder; Derouet, Mathieu F; Guo, Li; Karoubi, Golnaz; Silberberg, Amanda; Wong, Amy P; Waddell, Thomas K

    2017-03-01

    Cystic fibrosis (CF) is a fatal recessive genetic disorder caused by a mutation in the gene encoding CF transmembrane conductance regulator (CFTR) protein. Alteration in CFTR leads to thick airway mucus and bacterial infection. Cell therapy has been proposed for CFTR restoration, but efficacy has been limited by low engraftment levels. In our previous studies, we have shown that using a pre-conditioning regimen in combination with optimization of cell number and time of delivery, we could obtain greater bone marrow cell (BMC) retention in the lung. Here, we found that optimized delivery of wild-type (WT) BMC contributed to apical CFTR expression in airway epithelium and restoration of select ceramide species and fatty acids in CFTR(-/-) mice. Importantly, WT BMC delivery delayed Pseudomonas aeruginosa lung infection and increased survival of CFTR(-/-) recipients. Only WT BMCs had a beneficial effect beyond 6 months, suggesting a dual mechanism of BMC benefit: a non-specific effect early after cell delivery, possibly due to the recruitment of macrophages and neutrophils, and a late beneficial effect dependent on long-term CFTR expression. Taken together, our results suggest that BMC can improve overall lung function and may have potential therapeutic benefit for the treatment of CF.

  9. Differential downregulation of ACE2 by the spike proteins of severe acute respiratory syndrome coronavirus and human coronavirus NL63.

    PubMed

    Glowacka, Ilona; Bertram, Stephanie; Herzog, Petra; Pfefferle, Susanne; Steffen, Imke; Muench, Marcus O; Simmons, Graham; Hofmann, Heike; Kuri, Thomas; Weber, Friedemann; Eichler, Jutta; Drosten, Christian; Pöhlmann, Stefan

    2010-01-01

    The human coronaviruses (CoVs) severe acute respiratory syndrome (SARS)-CoV and NL63 employ angiotensin-converting enzyme 2 (ACE2) for cell entry. It was shown that recombinant SARS-CoV spike protein (SARS-S) downregulates ACE2 expression and thereby promotes lung injury. Whether NL63-S exerts a similar activity is yet unknown. We found that recombinant SARS-S bound to ACE2 and induced ACE2 shedding with higher efficiency than NL63-S. Shedding most likely accounted for the previously observed ACE2 downregulation but was dispensable for viral replication. Finally, SARS-CoV but not NL63 replicated efficiently in ACE2-positive Vero cells and reduced ACE2 expression, indicating robust receptor interference in the context of SARS-CoV but not NL63 infection.

  10. Differential renal effects of candesartan at high and ultra-high doses in diabetic mice–potential role of the ACE2/AT2R/Mas axis

    PubMed Central

    Callera, Glaucia E.; Antunes, Tayze T.; Correa, Jose W.; Moorman, Danielle; Gutsol, Alexey; He, Ying; Cat, Aurelie Nguyen Dinh; Briones, Ana M.; Montezano, Augusto C.; Burns, Kevin D.; Touyz, Rhian M.

    2016-01-01

    High doses of Ang II receptor (AT1R) blockers (ARBs) are renoprotective in diabetes. Underlying mechanisms remain unclear. We evaluated whether high/ultra-high doses of candesartan (ARB) up-regulate angiotensin-converting enzyme 2 (ACE2)/Ang II type 2 receptor (AT2R)/Mas receptor [protective axis of the of the renin–angiotensin system (RAS)] in diabetic mice. Systolic blood pressure (SBP), albuminuria and expression/activity of RAS components were assessed in diabetic db/db and control db/+ mice treated with increasing candesartan doses (intermediate, 1 mg/kg/d; high, 5 mg/kg/d; ultra-high, 25 and 75 mg/kg/d; 4 weeks). Lower doses candesartan did not influence SBP, but ultra-high doses reduced SBP in both groups. Plasma glucose and albuminuria were increased in db/db compared with db/+ mice. In diabetic mice treated with intermediate dose candesartan, renal tubular damage and albuminuria were ameliorated and expression of ACE2, AT2R and Mas and activity of ACE2 were increased, effects associated with reduced ERK1/2 phosphorylation, decreased fibrosis and renal protection. Ultra-high doses did not influence the ACE2/AT2R/Mas axis and promoted renal injury with increased renal ERK1/2 activation and exaggerated fibronectin expression in db/db mice. Our study demonstrates dose-related effects of candesartan in diabetic nephropathy: intermediate–high dose candesartan is renoprotective, whereas ultra-high dose candesartan induces renal damage. Molecular processes associated with these effects involve differential modulation of the ACE2/AT2R/Mas axis: intermediate–high dose candesartan up-regulating RAS protective components and attenuating pro-fibrotic processes, and ultra-high doses having opposite effects. These findings suggest novel mechanisms through the protective RAS axis, whereby candesartan may ameliorate diabetic nephropathy. Our findings also highlight potential injurious renal effects of ultra-high dose candesartan in diabetes. PMID:27612496

  11. Organic Anion–Transporting Polypeptide 1b2 (Oatp1b2) Is Important for the Hepatic Uptake of Unconjugated Bile Acids: Studies in Oatp1b2-Null Mice

    PubMed Central

    Csanaky, Iván L.; Lu, Hong; Zhang, Youcai; Ogura, Kenichiro; Choudhuri, Supratim; Klaassen, Curtis D.

    2011-01-01

    The organic anion–transporting polypeptide 1b family (Oatp1b2 in rodents and OATP1B1/1B3 in humans) is liver-specific and transports various chemicals into the liver. However, the role of the Oatp1b family in the hepatic uptake of bile acids (BAs) into the liver is unknown. Therefore, in Oatp1b2-null mice, the concentrations of BAs in plasma, liver, and bile were compared with wild-type (WT) mice. It was first determined that livers of the Oatp1b2-null mice were not compensated by altered expression of other hepatic transporters. However, the messenger RNA of Cyp7a1 was 70% lower in the Oatp1b2-null mice. Increased expression of fibroblast growth factor 15 in intestines of Oatp1b2-null mice might be responsible for decreased hepatic expression of Cyp7a1 in Oatp1b2-null mice. The hepatic concentration and biliary excretion of conjugated and unconjugated BAs were essentially the same in Oatp1b2-null and WT mice. The serum concentration of taurine-conjugated BAs was essentially the same in the two genotypes. In contrast, the serum concentrations of unconjugated BAs were 3–45 times higher in Oatp1b2-null than WT mice. After intravenous administration of cholate to Oatp1b2-null mice, its clearance was 50% lower than in WT mice, but the clearance of taurocholate was similar in the two genotypes. Conclusion This study indicates that Oatp1b2 has a major role in the hepatic uptake of unconjugated BAs. PMID:20949553

  12. Angiotensin-converting enzyme-2 (ACE2): comparative modeling of the active site, specificity requirements, and chloride dependence.

    PubMed

    Guy, Jodie L; Jackson, Richard M; Acharya, K Ravi; Sturrock, Edward D; Hooper, Nigel M; Turner, Anthony J

    2003-11-18

    Angiotensin-converting enzyme 2 (ACE2), a homologue of ACE, represents a new and potentially important target in cardio-renal disease. A model of the active site of ACE2, based on the crystal structure of testicular ACE, has been developed and indicates that the catalytic mechanism of ACE2 resembles that of ACE. Structural differences exist between the active site of ACE (dipeptidyl carboxypeptidase) and ACE2 (carboxypeptidase) that are responsible for the differences in specificity. The main differences occur in the ligand-binding pockets, particularly at the S2' subsite and in the binding of the peptide carboxy-terminus. The model explains why the classical ACE inhibitor lisinopril is unable to bind to ACE2. On the basis of the ability of ACE2 to cleave a variety of biologically active peptides, a consensus sequence of Pro-X-Pro-hydrophobic/basic for the protease specificity of ACE2 has been defined that is supported by the ACE2 model. The dipeptide, Pro-Phe, completely inhibits ACE2 activity at 180 microM with angiotensin II as the substrate. As with ACE, the chloride dependence of ACE2 is substrate-specific such that the hydrolysis of angiotensin I and the synthetic peptide substrate, Mca-APK(Dnp), are activated in the presence of chloride ions, whereas the cleavage of angiotensin II is inhibited. The ACE2 model is also suggestive of a possible mechanism for chloride activation. The structural insights provided by these analyses for the differences in inhibition pattern and substrate specificity among ACE and its homologue ACE2 and for the chloride dependence of ACE/ACE2 activity are valuable in understanding the function and regulation of ACE2.

  13. Sex difference in neural tube defects in p53-null mice is caused by differences in the complement of X not Y genes.

    PubMed

    Chen, Xuqi; Watkins, Rebecca; Delot, Emmanuele; Reliene, Ramune; Schiestl, Robert H; Burgoyne, Paul S; Arnold, Arthur P

    2008-02-01

    To shed light on the biological origins of sex differences in neural tube defects (NTDs), we examined Trp53-null C57BL/6 mouse embryos and neonates at 10.5 and 18.5 days post coitus (dpc) and at birth. We confirmed that female embryos show more NTDs than males. We also examined mice in which the testis-determining gene Sry is deleted from the Y chromosome but inserted onto an autosome as a transgene, producing XX and XY gonadal females and XX and XY gonadal males. At birth, Trp53 nullizygous mice were predominantly XY rather than XX, irrespective of gonadal type, showing that the sex difference in the lethal effect of Trp53 nullizygosity by postnatal day 1 is caused by differences in sex chromosome complement. At 10.5 dpc, the incidence of NTDs in Trp53-null progeny of XY* mice, among which the number of the X chromosomes varies independently of the presence or absence of a Y chromosome, was higher in mice with two copies of the X chromosome than in mice with a single copy. The presence of a Y chromosome had no protective effect, suggesting that sex differences in NTDs are caused by sex differences in the number of X chromosomes.

  14. Genome-Wide Screening of Alpha-Tocopherol Sensitive Genes in Heart Tissue from Alpha-Tocopherol Transfer Protein Null Mice (ATTP−/−)

    PubMed Central

    Vasu, Vihas T.; Hobson, Brad; Gohil, Kishorchandra; Cross, Carroll E.

    2009-01-01

    Alpha tocopherol transfer protein (ATTP) null mice (ATTP−/−) have a systemic deficiency of alpha-tocopherol (AT). The heart AT levels of ATTP−/− are <10% of those in ATTP+/+ mice. The genomic responses of heart to AT deficiency were determined in 3 months old male ATTP−/− mice and compared with their ATTP+/+ littermate controls using Affymetrix 430A 2.0 high density oligonucleotide arrays. Differential analysis of ~13,000 genes identified repression of genes related to immune system and activation of genes related to lipid metabolism and inflammation with no significant change in the expression of classical antioxidant genes (catalase, superoxide dismutase, glutathione peroxidase) in ATTP−/− as compared to ATTP+/+ mice. The present data identifies novel classes of AT sensitive genes in heart tissue. PMID:17382327

  15. FosB Null Mutant Mice Show Enhanced Methamphetamine Neurotoxicity: Potential Involvement of FosB in Intracellular Feedback Signaling and Astroglial Function

    PubMed Central

    Kuroda, Kumi O; Ornthanalai, Veravej G; Kato, Tadafumi; Murphy, Niall P

    2010-01-01

    Previous studies show that (1) two members of fos family transcription factors, c-Fos and FosB, are induced in frontal brain regions by methamphetamine; (2) null mutation of c-Fos exacerbates methamphetamine-induced neurotoxicity; and (3) null mutation of FosB enhances behavioral responses to cocaine. Here we sought a role of FosB in responses to methamphetamine by studying FosB null mutant (−/−) mice. After a 10 mg/kg methamphetamine injection, FosB(−/−) mice were more prone to self-injury. Concomitantly, the intracellular feedback regulators of Sprouty and Rad-Gem-Kir (RGK) family transcripts had lower expression profiles in the frontoparietal cortex and striatum of the FosB(−/−) mice. Three days after administration of four 10 mg/kg methamphetamine injections, the frontoparietal cortex and striatum of FosB(−/−) mice contained more degenerated neurons as determined by Fluoro-Jade B staining. The abundance of the small neutral amino acids, serine, alanine, and glycine, was lower and/or was poorly induced after methamphetamine administration in the frontoparietal cortex and striatum of FosB(−/−) mice. In addition, methamphetamine-treated FosB(−/−) frontoparietal and piriform cortices showed more extravasation of immunoglobulin, which is indicative of blood–brain barrier dysfunction. Methamphetamine-induced hyperthermia, brain dopamine content, and loss of tyrosine hydroxylase immunoreactivity in the striatum, however, were not different between genotypes. These data indicate that FosB is involved in thermoregulation-independent protective functions against methamphetamine neurotoxicity in postsynaptic neurons. Our findings suggest two possible mechanisms of FosB-mediated neuroprotection: one is induction of negative feedback regulation within postsynaptic neurons through Sprouty and RGK. Another is supporting astroglial function such as maintenance of the blood–brain barrier, and metabolism of serine and glycine, which are important

  16. Development of an autism severity score for mice using Nlgn4 null mutants as a construct-valid model of heritable monogenic autism.

    PubMed

    El-Kordi, Ahmed; Winkler, Daniela; Hammerschmidt, Kurt; Kästner, Anne; Krueger, Dilja; Ronnenberg, Anja; Ritter, Caroline; Jatho, Jasmin; Radyushkin, Konstantin; Bourgeron, Thomas; Fischer, Julia; Brose, Nils; Ehrenreich, Hannelore

    2013-08-15

    Autism is the short name of a complex and heterogeneous group of disorders (autism spectrum disorders, ASD) with several lead symptoms required for classification, including compromised social interaction, reduced verbal communication and stereotyped repetitive behaviors/restricted interests. The etiology of ASD is still unknown in most cases but monogenic heritable forms exist that have provided insights into ASD pathogenesis and have led to the notion of autism as a 'synapse disorder'. Among the most frequent monogenic causes of autism are loss-of-function mutations of the NLGN4X gene which encodes the synaptic cell adhesion protein neuroligin-4X (NLGN4X). We previously described autism-like behaviors in male Nlgn4 null mutant mice, including reduced social interaction and ultrasonic communication. Here, we extend the phenotypical characterization of Nlgn4 null mutant mice to both genders and add a series of additional autism-relevant behavioral readouts. We now report similar social interaction and ultrasonic communication deficits in females as in males. Furthermore, aggression, nest-building parameters, as well as self-grooming and circling as indicators of repetitive behaviors/stereotypies were explored in both genders. The construction of a gender-specific autism severity composite score for Nlgn4 mutant mice markedly diminishes population/sample heterogeneity typically obtained for single tests, resulting in p values of <0.00001 and a genotype predictability of 100% for male and of >83% for female mice. Taken together, these data underscore the similarity of phenotypical consequences of Nlgn4/NLGN4X loss-of-function in mouse and man, and emphasize the high relevance of Nlgn4 null mutant mice as an ASD model with both construct and face validity.

  17. FosB null mutant mice show enhanced methamphetamine neurotoxicity: potential involvement of FosB in intracellular feedback signaling and astroglial function.

    PubMed

    Kuroda, Kumi O; Ornthanalai, Veravej G; Kato, Tadafumi; Murphy, Niall P

    2010-02-01

    Previous studies show that (1) two members of fos family transcription factors, c-Fos and FosB, are induced in frontal brain regions by methamphetamine; (2) null mutation of c-Fos exacerbates methamphetamine-induced neurotoxicity; and (3) null mutation of FosB enhances behavioral responses to cocaine. Here we sought a role of FosB in responses to methamphetamine by studying FosB null mutant (-/-) mice. After a 10 mg/kg methamphetamine injection, FosB(-/-) mice were more prone to self-injury. Concomitantly, the intracellular feedback regulators of Sprouty and Rad-Gem-Kir (RGK) family transcripts had lower expression profiles in the frontoparietal cortex and striatum of the FosB(-/-) mice. Three days after administration of four 10 mg/kg methamphetamine injections, the frontoparietal cortex and striatum of FosB(-/-) mice contained more degenerated neurons as determined by Fluoro-Jade B staining. The abundance of the small neutral amino acids, serine, alanine, and glycine, was lower and/or was poorly induced after methamphetamine administration in the frontoparietal cortex and striatum of FosB(-/-) mice. In addition, methamphetamine-treated FosB(-/-) frontoparietal and piriform cortices showed more extravasation of immunoglobulin, which is indicative of blood-brain barrier dysfunction. Methamphetamine-induced hyperthermia, brain dopamine content, and loss of tyrosine hydroxylase immunoreactivity in the striatum, however, were not different between genotypes. These data indicate that FosB is involved in thermoregulation-independent protective functions against methamphetamine neurotoxicity in postsynaptic neurons. Our findings suggest two possible mechanisms of FosB-mediated neuroprotection: one is induction of negative feedback regulation within postsynaptic neurons through Sprouty and RGK. Another is supporting astroglial function such as maintenance of the blood-brain barrier, and metabolism of serine and glycine, which are important glial modulators of nerve cells.

  18. Role of Serratia marcescens ACE2 on diesel degradation and its influence on corrosion.

    PubMed

    Rajasekar, Aruliah; Babu, Thambidurai Ganesh; Pandian, Shunmugiah Thevar Karutha; Maruthamuthu, Sundaram; Palaniswamy, Narayanan; Rajendran, Annamalai

    2007-09-01

    A facultative anaerobic species Serratia marcescens ACE2 isolated from the corrosion products of diesel transporting pipeline in North West, India was identified by 16S rDNA sequence analysis. The role of Serratia marcesens ACE2 on biodegradation of diesel and its influence on the corrosion of API 5LX steel has been elucidated. The degrading strain ACE2 is involved in the process of corrosion of steel API 5LX and also utilizes the diesel as an organic source. The quantitative biodegradation efficiency (BE) of diesel was 58%, calculated by gas-chromatography-mass spectrum analysis. On the basis of gas-chromatography-mass spectrum (GC-MS), Fourier Transform infrared spectroscopy (FTIR) and X-ray diffractometer (XRD), the involvement of Serratia marcescens on degradation and corrosion has been investigated. This basic study will be useful for the development of new approaches for detection, monitoring and control of microbial corrosion.

  19. Bacterial Infection of Smad3/Rag2 Double-Null Mice with Transforming Growth Factor-β Dysregulation as a Model for Studying Inflammation-Associated Colon Cancer

    PubMed Central

    Maggio-Price, Lillian; Treuting, Piper; Bielefeldt-Ohmann, Helle; Seamons, Audrey; Drivdahl, Rolf; Zeng, Weiping; Lai, LapHin; Huycke, Mark; Phelps, Susan; Brabb, Thea; Iritani, Brian M.

    2009-01-01

    Alterations in genes encoding transforming growth factor-β-signaling components contribute to colon cancer in humans. Similarly, mice deficient in the transforming growth factor-β signaling molecule, Smad3, develop colon cancer, but only after a bacterial trigger occurs, resulting in chronic inflammation. To determine whether Smad3-null lymphocytes contribute to increased cancer susceptibility, we crossed Smad3-null mice with mice deficient in both B and T lymphocytes (Rag2−/− mice). Helicobacter-infected Smad3/Rag2-double knockout (DKO) mice had more diffuse inflammation and increased incidence of adenocarcinoma compared with Helicobacter-infected Smad3−/− or Rag2−/− mice alone. Adoptive transfer of WT CD4+CD25+ T-regulatory cells provided significant protection of Smad3/Rag2-DKO from bacterial-induced typhlocolitis, dysplasia, and tumor development, whereas Smad3−/− T-regulatory cells provided no protection. Immunohistochemistry, real-time reverse transcriptase-polymerase chain reaction, and Western blot analyses of colonic tissues from Smad3/Rag2-DKO mice 1 week after Helicobacter infection revealed an influx of macrophages, enhanced nuclear factor-κB activation, increased BclXL/Bcl-2 expression, increased c-Myc expression, accentuated epithelial cell proliferation, and up-regulated IFN-γ, IL-1α, TNF-α, IL-1β, and IL-6 transcription levels. These results suggest that the loss of Smad3 increases susceptibility to colon cancer by at least two mechanisms: deficient T-regulatory cell function, which leads to excessive inflammation after a bacterial trigger; and increased expression of proinflammatory cytokines, enhanced nuclear factor-κB activation, and increased expression of both pro-oncogenic and anti-apoptotic proteins that result in increased cell proliferation/survival of epithelial cells in colonic tissues. PMID:19119184

  20. Improved function and proliferation of adult human beta cells engrafted in diabetic immunodeficient NOD-scid IL2rγnull mice treated with alogliptin

    PubMed Central

    Jurczyk, Agata; diIorio, Philip; Brostowin, Dean; Leehy, Linda; Yang, Chaoxing; Urano, Fumihiko; Harlan, David M; Shultz, Leonard D; Greiner, Dale L; Bortell, Rita

    2013-01-01

    Purpose Dipeptidyl-peptidase-4 (DPP-4) inhibitors are known to increase insulin secretion and beta cell proliferation in rodents. To investigate the effects on human beta cells in vivo, we utilize immunodeficient mice transplanted with human islets. The study goal was to determine the efficacy of alogliptin, a DPP-4 inhibitor, to enhance human beta cell function and proliferation in an in vivo context using diabetic immunodeficient mice engrafted with human pancreatic islets. Methods Streptozotocin-induced diabetic NOD-scid IL2rγnull (NSG) mice were transplanted with adult human islets in three separate trials. Transplanted mice were treated daily by gavage with alogliptin (30 mg/kg/day) or vehicle control. Islet graft function was compared using glucose tolerance tests and non-fasting plasma levels of human insulin and C-peptide; beta cell proliferation was determined by bromodeoxyuridine (BrdU) incorporation. Results Glucose tolerance tests were significantly improved by alogliptin treatment for mice transplanted with islets from two of the three human islet donors. Islet-engrafted mice treated with alogliptin also had significantly higher plasma levels of human insulin and C-peptide compared to vehicle controls. The percentage of insulin+BrdU+ cells in human islet grafts from alogliptin-treated mice was approximately 10-fold more than from vehicle control mice, consistent with a significant increase in human beta cell proliferation. Conclusion Human islet-engrafted immunodeficient mice treated with alogliptin show improved human insulin secretion and beta cell proliferation compared to control mice engrafted with the same donor islets. Immunodeficient mice transplanted with human islets provide a useful model to interrogate potential therapies to improve human islet function and survival in vivo. PMID:24376359

  1. Comparison of germ cell mutagenicity in male CYP2E1-null and wild-type mice treated with acrylamide: evidence supporting a glycidamide-mediated effect.

    PubMed

    Ghanayem, B I; Witt, K L; El-Hadri, L; Hoffler, U; Kissling, G E; Shelby, M D; Bishop, J B

    2005-01-01

    Acrylamide is an animal carcinogen and probable human carcinogen present in appreciable amounts in heated carbohydrate-rich foodstuffs. It is also a germ cell mutagen, inducing dominant lethal mutations and heritable chromosomal translocations in postmeiotic sperm of treated mice. Acrylamide's affinity for male germ cells has sometimes been overlooked in assessing its toxicity and defining human health risks. Previous investigations of acrylamide's germ cell activity in mice showed stronger effects after repeated administration of low doses compared with a single high dose, suggesting the possible involvement of a stable metabolite. A key oxidative metabolite of acrylamide is the epoxide glycidamide, generated by cytochrome P4502E1 (CYP2E1). To explore the role of CYP2E1 metabolism in the germ cell mutagenicity of acrylamide, CYP2E1-null and wild-type male mice were treated by intraperitoneal injection with 0, 12.5, 25, or 50 mg acrylamide (5 ml saline)(-1) kg(-1) day(-1) for 5 consecutive days. At defined times after exposure, males were mated to untreated B6C3F1 females. Females were killed in late gestation and uterine contents were examined. Dose-related increases in resorption moles (chromosomally aberrant embryos) and decreases in the numbers of pregnant females and the proportion of living fetuses were seen in females mated to acrylamide-treated wild-type mice. No changes in any fertility parameters were seen in females mated to acrylamide-treated CYP2E1-null mice. Our results constitute the first unequivocal demonstration that acrylamide-induced germ cell mutations in male mice require CYP2E1-mediated epoxidation of acrylamide. Thus, CYP2E1 polymorphisms in human populations, resulting in variable enzyme metabolic activities, may produce differential susceptibilities to acrylamide toxicities.

  2. Dystrophin and utrophin expression require sarcospan: loss of α7 integrin exacerbates a newly discovered muscle phenotype in sarcospan-null mice.

    PubMed

    Marshall, Jamie L; Chou, Eric; Oh, Jennifer; Kwok, Allan; Burkin, Dean J; Crosbie-Watson, Rachelle H

    2012-10-15

    Sarcospan (SSPN) is a core component of the major adhesion complexes in skeletal muscle, the dystrophin- and utrophin (Utr)-glycoprotein complexes (DGC and UGC). We performed a rigorous analysis of SSPN-null mice and discovered that loss of SSPN decreased DGC and UGC abundance, leading to impaired laminin-binding activity and susceptibility to eccentric contraction-induced injury in skeletal muscle. We show that loss of SSPN increased levels of α7β1 integrin. To genetically test whether integrin compensates for the loss of DGC and UGC function in SSPN-nulls, we generated mice lacking both SSPN and α7 integrin (DKO, double knockout). Muscle regeneration, sarcolemma integrity and fibrosis were exacerbated in DKO mice and were remarkably similar to muscle from Duchenne muscular dystrophy (DMD) patients, suggesting that secondary loss of integrin contributes significantly to pathogenesis. Expression of the DGC and UGC, laminin binding and Akt signaling were negatively impacted in DKO muscle, resulting in severely diminished specific force properties. We demonstrate that SSPN is a necessary component of dystrophin and Utr function and that SSPN modulation of integrin signaling is required for extracellular matrix attachment and muscle force development.

  3. Hepatic effects of repeated oral administration of diclofenac to hepatic cytochrome P450 reductase null (HRN™) and wild-type mice.

    PubMed

    Akingbasote, James A; Foster, Alison J; Wilson, Ian; Sarda, Sunil; Jones, Huw B; Kenna, J Gerry

    2016-04-01

    Hepatic NADPH-cytochrome P450 oxidoreductase null (HRN™) mice exhibit normal hepatic and extrahepatic biotransformation enzyme activities when compared to wild-type (WT) mice, but express no functional hepatic cytochrome P450 activities. When incubated in vitro with [(14)C]-diclofenac, liver microsomes from WT mice exhibited extensive biotransformation to oxidative and glucuronide metabolites and covalent binding to proteins was also observed. In contrast, whereas glucuronide conjugates and a quinone-imine metabolite were formed when [(14)C]-diclofenac was incubated with HRN™ mouse liver, only small quantities of P450-derived oxidative metabolites were produced in these samples and covalent binding to proteins was not observed. Livers from vehicle-treated HRN™ mice exhibited enhanced lipid accumulation, bile duct proliferation, hepatocellular degeneration and necrosis and inflammatory cell infiltration, which were not present in livers from WT mice. Elevated liver-derived alanine aminotransferase, glutamate dehydrogenase and alkaline phosphatase activities were also observed in plasma from HRN™ mice. When treated orally with diclofenac for 7 days, at 30 mg/kg/day, the severities of the abnormal liver histopathology and plasma liver enzyme findings in HRN™ mice were reduced markedly. Oral diclofenac administration did not alter the liver histopathology or elevate plasma enzyme activities of WT mice. These findings indicate that HRN™ mice are valuable for exploration of the role played by hepatic P450s in drug biotransformation, but poorly suited to investigations of drug-induced liver toxicity. Nevertheless, studies in HRN™ mice could provide novel insights into the role played by inflammation in liver injury and may aid the evaluation of new strategies for its treatment.

  4. Activation of the ACE2/Ang-(1-7)/Mas pathway reduces oxygen-glucose deprivation induced tissue swelling, ROS production, and cell death in mouse brain with angiotensin II overproduction

    PubMed Central

    Zheng, Jiaolin; Li, Guangze; Chen, Shuzhen; Chen, Ji; Buck, Joshua; Zhu, Yulan; Xia, Huijing; Lazartigues, Eric; Chen, Yanfang; Olson, James E.

    2014-01-01

    We previously demonstrated that mice which overexpress human renin and angiotensinogen (R+A+) show enhanced cerebral damage in both in vivo and in vitro experimental ischemia models. Angiotensin converting enzyme 2 (ACE2) counteracts the effects of angiotensin (Ang-II) by transforming it into Ang-(1-7), thus reducing the ligand for the AT1 receptor and increasing stimulation of the Mas receptor. Triple transgenic mice, SARA, which specifically overexpress ACE2 in neurons of R+A+ mice were used to study the role of ACE2 in ischemic stroke using oxygen and glucose deprivation (OGD) of brain slices as an in vitro model. We examined tissue swelling, the production of reactive oxygen species (ROS), and cell death in cerebral cortex (CX) and the hippocampal CA1 region during OGD. Expression levels of NADPH oxidase isoforms, Nox2 and Nox4 were measured using western blots. Results show that SARA mice and R+A+ mice treated with the Mas receptor agonist Ang-(1-7) had less swelling, cell death, and ROS production in CX and CA1 areas compared to those in R+A+ animals. Treatment of slices from SARA mice with the Mas antagonist A779 eliminated this protection. Finally, western blots revealed less Nox2 and Nox4 expression in SARA mice compared with R+A+ mice both before and after OGD. We suggest that reduced brain swelling and cell death observed in SARA animals exposed to OGD results from diminished ROS production coupled with lower expression of NADPH oxidases. Thus, the ACE2/Ang-(1-7)/Mas receptor pathway plays a protective role in brain ischemic damage by counteracting the detrimental effects of Ang-II-induced ROS production. PMID:24814023

  5. Engraftment of human HSCs in nonirradiated newborn NOD-scid IL2rγnull mice is enhanced by transgenic expression of membrane-bound human SCF

    PubMed Central

    Racki, Waldemar J.; Leif, Jean; Burzenski, Lisa; Hosur, Vishnu; Wetmore, Amber; Gott, Bruce; Herlihy, Mary; Ignotz, Ronald; Dunn, Raymond; Shultz, Leonard D.; Greiner, Dale L.

    2012-01-01

    Immunodeficient mice engrafted with human HSCs support multidisciplinary translational experimentation, including the study of human hematopoiesis. Heightened levels of human HSC engraftment are observed in immunodeficient mice expressing mutations in the IL2-receptor common γ chain (IL2rg) gene, including NOD-scid IL2rγnull (NSG) mice. Engraftment of human HSC requires preconditioning of immunodeficient recipients, usually with irradiation. Such preconditioning increases the expression of stem cell factor (SCF), which is critical for HSC engraftment, proliferation, and survival. We hypothesized that transgenic expression of human membrane-bound stem cell factor Tg(hu-mSCF)] would increase levels of human HSC engraftment in nonirradiated NSG mice and eliminate complications associated with irradiation. Surprisingly, detectable levels of human CD45+ cell chimerism were observed after transplantation of cord blood–derived human HSCs into nonirradiated adult as well as newborn NSG mice. However, transgenic expression of human mSCF enabled heightened levels of human hematopoietic cell chimerism in the absence of irradiation. Moreover, nonirradiated NSG-Tg(hu-mSCF) mice engrafted as newborns with human HSCs rejected human skin grafts from a histoincompatible donor, indicating the development of a functional human immune system. These data provide a new immunodeficient mouse model that does not require irradiation preconditioning for human HSC engraftment and immune system development. PMID:22246028

  6. Ly-2+ T cell enlargement and null cell proliferation occur at the onset of splenomegaly and autoantibody production in New Zealand Black mice.

    PubMed

    Manohar, V; Brown, E M; Leiserson, W M; Edison, L J; Chused, T M

    1984-12-01

    Splenic T and B lymphocyte and null cell populations were analyzed in NZB mice as autoimmune disease developed during the first year of life. In the B lymphocytes, a progressive shift occurred from the surface IgD bright subset to the surface IgM bright subset. There was a slight increase in the ratio of Ly-2- to Ly-2+ T cells. Splenomegaly was not detected until after 40 wk of age and was primarily due to an increase in the number of null cells. This change was accompanied by an increase in the size, as determined by narrow-angle forward light scatter, of the Ly-2+ but not the Ly-2- T cells, an elevation of IgG-containing plasma cells, and the appearance of anti-erythrocyte autoantibody. The splenic B cell subset distribution and the enlargement of the Ly-2+ T cells were reflected in the peripheral blood, whereas the T cell subset ratio was not. The B cell subset alteration did not correlate with any of the other changes observed. Statistical associations were found between the ratios of T cell subsets, the enlargement of the Ly-2+ T cells, and the increased number of null cells, suggesting a linkage among those late changes that immediately precede the development of overt autoimmune disease.

  7. Human intestine luminal ACE2 and amino acid transporter expression increased by ACE-inhibitors.

    PubMed

    Vuille-dit-Bille, Raphael N; Camargo, Simone M; Emmenegger, Luca; Sasse, Tom; Kummer, Eva; Jando, Julia; Hamie, Qeumars M; Meier, Chantal F; Hunziker, Schirin; Forras-Kaufmann, Zsofia; Kuyumcu, Sena; Fox, Mark; Schwizer, Werner; Fried, Michael; Lindenmeyer, Maja; Götze, Oliver; Verrey, François

    2015-04-01

    Sodium-dependent neutral amino acid transporter B(0)AT1 (SLC6A19) and imino acid (proline) transporter SIT1 (SLC6A20) are expressed at the luminal membrane of small intestine enterocytes and proximal tubule kidney cells where they exert key functions for amino acid (re)absorption as documented by their role in Hartnup disorder and iminoglycinuria, respectively. Expression of B(0)AT1 was shown in rodent intestine to depend on the presence of the carboxypeptidase angiotensin-converting enzyme 2 (ACE2). This enzyme belongs to the renin-angiotensin system and its expression is induced by treatment with ACE-inhibitors (ACEIs) or angiotensin II AT1 receptor blockers (ARBs) in many rodent tissues. We show here in the Xenopus laevis oocyte expression system that human ACE2 also functionally interacts with SIT1. To investigate in human intestine the potential effect of ACEIs or ARBs on ACE2, we analysed intestinal biopsies taken during routine gastroduodenoscopy and ileocolonoscopy from 46 patients of which 9 were under ACEI and 13 ARB treatment. Analysis of transcript expression by real-time PCR and of proteins by immunofluorescence showed a co-localization of SIT1 and B(0)AT1 with ACE2 in the brush-border membrane of human small intestine enterocytes and a distinct axial expression pattern of the tested gene products along the intestine. Patients treated with ACEIs displayed in comparison with untreated controls increased intestinal mRNA levels of ACE2, peptide transporter PEPT1 (SLC15A1) and AA transporters B(0)AT1 and PAT1 (SLC36A1). This study unravels in human intestine the localization and distribution of intestinal transporters involved in amino acid absorption and suggests that ACEIs impact on their expression.

  8. Diminazene aceturate enhances ACE2 activity and attenuates ischemia-induced cardiac pathophysiology

    PubMed Central

    Qi, YanFei; Zhang, Juan; Cole-Jeffrey, Colleen T; Shenoy, Vinayak; Espejo, Andrew; Hanna, Mina; Song, Chunjuan; Pepine, Carl J; Katovich, Michael J; Raizada, Mohan K

    2013-01-01

    Angiotensin-converting enzyme 2 (ACE2) plays a critical role against myocardial infarction (MI). We hypothesized that activation of intrinsic ACE2 would be protective against ischemia-induced cardiac pathophysiology. Diminazine aceturate (DIZE), a small molecule ACE2 activator has been used to evaluate this hypothesis. DIZE (15 mg/kg/day, s.c.) was injected two days prior to MI surgery and continued throughout the study-period. MI rats showed a 62% decrease in fractional shortening (FS,%) [control (Con): 51.1 ± 3.2; DIZE alone (D) : 52.1 ± 3.2; MI (M): 19.1± 3.0], a 55% decrease in contractility (dP/dtmax mmHg/s) (Con: 9480 ± 425.3; D: 9585 ± 597.4; M: 4251 ± 657.7), and a 27% increase in ventricular hypertrophy [VH, mg/mm (Con: 26.5 ± 1.5; D: 26.9 ± 1.4; M: 33.4± 1.1)]. DIZE attenuated the MI-induced decrease in FS by 89%, improved dP/dtmax by 92%, and reversed VH by 18%. MI also significantly increased ACE and angiotensin type 1 receptor levels while decreased ACE2 activity by 40% (Con: 246.2 ± 25.1; D: 254.2 ± 20.6; M: 148.9 ± 29.2, RFU/min), which was reversed by DIZE treatment. Thus, DIZE treatment decreased the infarct area, attenuated LV remodeling post-MI and restored normal balance of the cardiac renin angiotensin system. Additionally, DIZE treatment increased circulating endothelial progenitor cells, increased engraftment of cardiac progenitor cells and decreased inflammatory cells in peri-infarct cardiac regions. All of the beneficial effects associated with DIZE treatment were abolished by C-16, an ACE2 inhibitor. Collectively, DIZE and DIZE-like small molecules may represent promising new therapeutic agents for MI. PMID:23959549

  9. Role of angiotensin-converting enzyme 2 (ACE2) in diabetic cardiovascular complications.

    PubMed

    Patel, Vaibhav B; Parajuli, Nirmal; Oudit, Gavin Y

    2014-04-01

    Diabetes mellitus results in severe cardiovascular complications, and heart disease and failure remain the major causes of death in patients with diabetes. Given the increasing global tide of obesity and diabetes, the clinical burden of diabetes-induced cardiovascular disease is reaching epidemic proportions. Therefore urgent actions are needed to stem the tide of diabetes which entails new prevention and treatment tools. Clinical and pharmacological studies have demonstrated that AngII (angiotensin II), the major effector peptide of the RAS (renin-angiotensin system), is a critical promoter of insulin resistance and diabetes mellitus. The role of RAS and AngII has been implicated in the progression of diabetic cardiovascular complications and AT1R (AngII type 1 receptor) blockers and ACE (angiotensin-converting enzyme) inhibitors have shown clinical benefits. ACE2, the recently discovered homologue of ACE, is a monocarboxypeptidase which converts AngII into Ang-(1-7) [angiotensin-(1-7)] which, by virtue of its actions on the MasR (Mas receptor), opposes the effects of AngII. In animal models of diabetes, an early increase in ACE2 expression and activity occurs, whereas ACE2 mRNA and protein levels have been found to decrease in older STZ (streptozotocin)-induced diabetic rats. Using the Akita mouse model of Type 1 diabetes, we have recently shown that loss of ACE2 disrupts the balance of the RAS in a diabetic state and leads to AngII/AT1R-dependent systolic dysfunction and impaired vascular function. In the present review, we will discuss the role of the RAS in the pathophysiology and treatment of diabetes and its complications with particular emphasis on potential benefits of the ACE2/Ang-(1-7)/MasR axis activation.

  10. Rescue of motor coordination by Purkinje cell-targeted restoration of Kv3.3 channels in Kcnc3-null mice requires Kcnc1.

    PubMed

    Hurlock, Edward C; Bose, Mitali; Pierce, Ganon; Joho, Rolf H

    2009-12-16

    The role of cerebellar Kv3.1 and Kv3.3 channels in motor coordination was examined with an emphasis on the deep cerebellar nuclei (DCN). Kv3 channel subunits encoded by Kcnc genes are distinguished by rapid activation and deactivation kinetics that support high-frequency, narrow action potential firing. Previously we reported that increased lateral deviation while ambulating and slips while traversing a narrow beam of ataxic Kcnc3-null mice were corrected by restoration of Kv3.3 channels specifically to Purkinje cells, whereas Kcnc3-mutant mice additionally lacking one Kcnc1 allele were partially rescued. Here, we report mice lacking all Kcnc1 and Kcnc3 alleles exhibit no such rescue. For Purkinje cell output to reach the rest of the brain it must be conveyed by neurons of the DCN or vestibular nuclei. As Kcnc1, but not Kcnc3, alleles are lost, mutant mice exhibit increasing gait ataxia accompanied by spike broadening and deceleration in DCN neurons, suggesting the facet of coordination rescued by Purkinje-cell-restricted Kv3.3 restoration in mice lacking just Kcnc3 is hypermetria, while gait ataxia emerges when additionally Kcnc1 alleles are lost. Thus, fast repolarization in Purkinje cells appears important for normal movement velocity, whereas DCN neurons are a prime candidate locus where fast repolarization is necessary for normal gait patterning.

  11. Oral administration of L-mR18L, a single domain cationic amphipathic helical peptide, inhibits lesion formation in ApoE null mice.

    PubMed

    Handattu, Shaila P; Datta, Geeta; Epand, Richard M; Epand, Raquel F; Palgunachari, Mayakonda N; Mishra, Vinod K; Monroe, Candyce E; Keenum, Tamara D; Chaddha, Manjula; Anantharamaiah, G M; Garber, David W

    2010-12-01

    We have shown that Ac-hE18A-NH₂, a dual-domain cationic apolipoprotein-mimetic peptide, reduces plasma cholesterol levels in dyslipidemic mice. Two single-domain cationic peptides based on the lytic class L peptide 18L were developed to test the hypothesis that a single-domain cationic amphipathic peptide can reduce atherosclerosis in apolipoprotein (apo)E null mice when orally administered. To incorporate anti-inflammatory properties, aromatic residues were clustered in the nonpolar face similar to peptide 4F, resulting in modified 18L (m18L). To reduce lytic properties, the Lys residues of 18L were replaced with Arg with the resulting peptide called modified R18L (mR18L). Biophysical studies showed that mR18L had stronger interactions with lipids than did m18L. Peptide mR18L was also more effective than m18L in promoting LDL uptake by HepG2 cells. ApoE null mice received normal chow or chow containing m18L or mR18L for six weeks. A significant reduction in plasma cholesterol and aortic sinus lesion area was seen only in the mR18L group. Plasma from mice administered mR18L, unlike those from the control and m18L groups, did not enhance monocyte adhesion to endothelial cells. Thus oral administration of mR18L reduces plasma cholesterol and lesion formation and inhibits monocyte adhesion.

  12. Expression in SPARC-null mice of collagen type I lacking the globular domain of the α1(I) N-propeptide results in abdominal hernias and loss of dermal collagen.

    PubMed

    Card, Lauren; Henderson, Nikki; Zhang, Yuhua; Bornstein, Paul; Bradshaw, Amy D

    2010-09-01

    The sequence encoding the N-propeptide of collagen I is characterized by significant conservation of amino acids across species; however, the function of the N-propeptide remains poorly defined. Studies in vitro have suggested that one activity of this propeptide might be to act as a feedback inhibitor of collagen I synthesis. To determine whether the N-propeptide contributed to decreased collagen content in SPARC-null mice, mice carrying a deletion of exon 2, which encodes the globular domain of the N-propeptide of collagen I, were crossed to SPARC-null animals. Mice lacking SPARC and expressing collagen I without the globular domain of the N-propeptide were viable and fertile. However, a significant number of animals developed abdominal hernias within the first 2 months of life with an approximate 20% penetrance (~35% of males). The dermis of SPARC-null/exon 2-deleted mice was thinner and contained fewer large collagen fibers in comparison with wild-type or in either single transgenic animal. The average collagen fibril diameter of exon 2-deleted mice did not significantly differ from wild-type mice (WT: 87.9 nm versus exon 2-deleted: 88.2 nm), whereas SPARC-null/exon 2-deleted fibrils were smaller than that of SPARC-null dermis (SPARC-null: 60.2 nm, SPARC-null/exon 2-deleted: 40.8 nm). As measured by hydroxyproline analysis, double transgenic skin biopsies contained significantly less collagen than those of wild-type, those of exon 2-deleted, and those of SPARC-null biopsies. Acetic acid extraction of collagen from skin biopsies revealed an increase in the proportion of soluble collagen in the SPARC-null/exon 2-deleted mice. These results support a function of the N-propeptide of collagen I in facilitating incorporation and stabilization of collagen I into the insoluble ECM and argue against a primary function of the N-propeptide as a negative regulator of collagen synthesis.

  13. NOD-scid IL2R γnull mice engrafted with human peripheral blood mononuclear cells as a model to test therapeutics targeting human signaling pathways

    PubMed Central

    2013-01-01

    Background Animal models of human inflammatory diseases have limited predictive quality for human clinical trials for various reasons including species specific activation mechanisms and the immunological background of the animals which markedly differs from the genetically heterogeneous and often aged patient population. Objective Development of an animal model allowing for testing therapeutics targeting pathways involved in the development of Atopic Dermatitis (AD) with better translatability to the patient. Methods NOD-scid IL2R γnull mice engrafted with human peripheral blood mononuclear cells (hPBMC) derived from patients suffering from AD and healthy volunteers were treated with IL-4 and the antagonistic IL-4 variant R121/Y124D (Pitrakinra). Levels of human (h)IgE, amount of B-, T- and plasma- cells and ratio of CD4 : CD8 positive cells served as read out for induction and inhibition of cell proliferation and hIgE secretion. Results were compared to in vitro analysis. Results hIgE secretion was induced by IL-4 and inhibited by the IL-4 antagonist Pitrakinra in vivo when formulated with methylcellulose. B-cells proliferated in response to IL-4 in vivo; the effect was abrogated by Pitrakinra. IL-4 shifted CD4 : CD8 ratios in vitro and in vivo when hPBMC derived from healthy volunteers were used. Pitrakinra reversed the effect. Human PBMC derived from patients with AD remained inert and engrafted mice reflected the individual responses observed in vitro. Conclusion NOD-scid IL2R γnull mice engrafted with human PBMC reflect the immunological history of the donors and provide a complementary tool to in vitro studies. Thus, studies in this model might provide data with better translatability from bench to bedside. PMID:23294516

  14. Timed conditional null of connexin26 in mice reveals temporary requirements of connexin26 in key cochlear developmental events before the onset of hearing.

    PubMed

    Chang, Qing; Tang, Wenxue; Kim, Yeunjung; Lin, Xi

    2015-01-01

    Mutations in the Gjb2 gene, which encodes a gap junction protein connexin26 (Cx26), are the most prevalent form of hereditary deafness in humans and represent about half of non-syndromic congenital deafness cases in many ethnic populations. Cochlear potassium (K+) recycling in mature cochlea is required for normal hearing. It is thought that gap junctions are essential for K+ recycling and that Gjb2 mutations cause Gjb2-associated deafness by disrupting K+ recycling in mature cochlea. Here we present evidence showing that Gjb2 is required for normal development of the neonatal organ of Corti prior to the onset of the hearing in mice. In the conditional Gjb2 null (cCx26 null) mice, ribbon synapses in inner hair cells remained poorly developed, the afferent type I fibers failed to finish the refinement process to form convergent innervation to individual inner hair cells. The spontaneous depolarizing activities in the supporting cells, which normally diminish in the wild type cochleae after postnatal day 8 (P8), remained strong in the cochlea after P8 in the mutant mice. Furthermore, the deafness phenotype was readily generated only if the Cx26 expression in the organ of Corti was significantly reduced before P6. Similar amount of Cx26 reduction in more mature cochleae had a much weaker effect in damaging the hearing sensitivity. Our findings indicated that Cx26 plays essential roles in the maturation process of the organ of Corti prior to the establishment of high K+ in the endolymph and the onset of hearing. These results suggest that successful treatment of Cx26 deafness requires early intervention before the cochlea fully matures.

  15. Identification of novel toxicity-associated metabolites by metabolomics and mass isotopomer analysis of acetaminophen metabolism in wild-type and Cyp2e1-null mice.

    PubMed

    Chen, Chi; Krausz, Kristopher W; Idle, Jeffrey R; Gonzalez, Frank J

    2008-02-22

    CYP2E1 is recognized as the most important enzyme for initiation of acetaminophen (APAP)-induced toxicity. In this study, the resistance of Cyp2e1-null mice to APAP treatment was confirmed by comparing serum aminotransferase activities and blood urea nitrogen levels in wild-type and Cyp2e1-null mice. However, unexpectedly, profiling of major known APAP metabolites in urine and serum revealed that the contribution of CYP2E1 to APAP metabolism decreased with increasing APAP doses administered. Measurement of hepatic glutathione and hydrogen peroxide levels exposed the importance of oxidative stress in determining the consequence of APAP overdose. Subsequent metabolomic analysis was capable of constructing a principal components analysis (PCA) model that delineated a relationship between urinary metabolomes and the responses to APAP treatment. Urinary ions high in wild-type mice treated with 400 mg/kg APAP were elucidated as 3-methoxy-APAP glucuronide (VII) and three novel APAP metabolites, including S-(5-acetylamino-2-hydroxyphenyl)mercaptopyruvic acid (VI, formed by a Cys-APAP transamination reaction in kidney), 3,3'-biacetaminophen (VIII, an APAP dimer), and a benzothiazine compound (IX, originated from deacetylated APAP), through mass isotopomer analysis, accurate mass measurement, tandem mass spectrometry fragmentation, in vitro reactions, and chemical treatments. Dose-, time-, and genotype-dependent appearance of these minor APAP metabolites implied their association with the APAP-induced toxicity and potential biomarker application. Overall, the oxidative stress elicited by CYP2E1-mediated APAP metabolism might significantly contribute to APAP-induced toxicity. The combination of genetically modified animal models, mass isotopomer analysis, and metabolomics provides a powerful and efficient technical platform to characterize APAP-induced toxicity through identifying novel biomarkers and unraveling novel mechanisms.

  16. Acute intermittent hypoxia-induced expression of brain-derived neurotrophic factor is disrupted in the brainstem of methyl-CpG-binding protein 2 null mice.

    PubMed

    Vermehren-Schmaedick, A; Jenkins, V K; Knopp, S J; Balkowiec, A; Bissonnette, J M

    2012-03-29

    Rett syndrome is a neurodevelopmental disorder caused by loss-of-function mutations in the gene encoding the transcription factor methyl-CpG-binding protein 2 (MeCP2). One of its targets is the gene encoding brain-derived neurotrophic factor (bdnf). In vitro studies using cultured neurons have produced conflicting results with respect to the role of MeCP2 in BDNF expression. Acute intermittent hypoxia (AIH) induces plasticity in the respiratory system characterized by long-term facilitation of phrenic nerve amplitude. This paradigm induces an increase in BDNF protein. We hypothesized that AIH leads to augmentation of BDNF transcription in respiratory-related areas of the brainstem and that MeCP2 is necessary for this process. Wild-type and mecp2 null (mecp2(-/y)) mice were subjected to three 5-min episodes of exposure to 8% O(2)/4% CO(2)/88% N(2), delivered at 5-min intervals. Normoxia control wild-type and mecp2 null mice were exposed to room air for the total length of time, that is, 30 min. Following a recovery in room air, the pons and medulla were rapidly removed. Expression of BDNF protein and transcripts were determined by ELISA and quantitative PCR, respectively. AIH induced a significant increase in BDNF protein in the pons and medulla, and in mRNA transcript levels in the pons of wild-type animals. In contrast, there were no significant changes in either BDNF protein or transcripts in the pons or medulla of mice lacking MeCP2. The results indicate that MeCP2 is required for regulation of BDNF expression by acute intermittent hypoxia in vivo.

  17. North Atlantic Aerosol Properties and Direct Radiative Effects: Key Results from TARFOX and ACE-2

    NASA Technical Reports Server (NTRS)

    Russell, P. B.; Livingston, J. M.; Schmid, B.; Bergstrom, R. A.; Hignett, P.; Hobbs, P. V.; Durkee, P. A.; Condon, Estelle (Technical Monitor)

    1998-01-01

    Aerosol effects on atmospheric radiative fluxes provide a forcing function that can change the climate in potentially significant ways. This aerosol radiative Forcing is a major source of uncertainty in understanding the observed climate change of the past century and in predicting, future climate. To help reduce this uncertainty, the International Global Atmospheric Chemistry Project (IGAC) has endorsed a series of multiplatform aerosol field campaigns. The Tropospheric Aerosol Radiative Forcing Observational Experiment (TARFOX) and the second Aerosol Characterization Experiment (ACE-2) were the first IGAC campaigns to address the impact of anthropogenic aerosols. Both TARFOX and ACE-2 gathered extensive data sets on aerosol properties and radiative effects. TARFOX focused on the urban-industrial haze plume flowing from the eastern United States over the western Atlantic Ocean, whereas ACE-2 studied aerosols carried over the eastern Atlantic from both European urban/industrial and African mineral sources. These aerosols often have a marked influence on the top-of-atmosphere radiances measured by satellites, as illustrated in Figure 1. Shown there are contours of aerosol optical depth derived from radiances measured by the AVHRR sensor on the NOAA-11 satellite. The contours readily show that aerosols originating in North America, Europe, and Africa impact the radiative properties of air over the North Atlantic. However, the accurate derivation of flux chances, or radiative forcing, from the satellite-measured radiances or 'etrieved optical depths remains a difficult challenge. In this paper we summarize key Initial results from TARFOX and, to a lesser extent ACE-2, with a focus on those results that allow an improved assessment of the flux changes caused by North Atlantic aerosols at middle and high latitudes.

  18. North Atlantic Aerosol Properties and Direct Radiative Effects: Key Results from TARFOX and ACE-2

    NASA Technical Reports Server (NTRS)

    Russell, P. B.; Livingston, J. M.; Schmid, B.; Bergstrom, Robert A.; Hignett, P.; Hobbs, P. V.; Durkee, P. A.

    2000-01-01

    Aerosol effects on atmospheric radiative fluxes provide a forcing function that can change the climate In potentially significant ways. This aerosol radiative forcing is a major source of uncertainty in understanding the observed climate change of the past century and in predicting future climate. To help reduce this uncertainty, the International Global Atmospheric Chemistry Project (IGAC) has endorsed a series of multiplatform aerosol field campaigns. The Tropospheric Aerosol Radiative Forcing Observational Experiment (TARFOX) and the second Aerosol Characterization Experiment (ACE-2) were the first IGAC campaigns to address the impact of anthropogenic aerosols, Both TARFOX and ACE-2 gathered extensive data sets on aerosol properties and radiative effects, TARFOX focused on the urban-industrial haze plume flowing from the eastern United States over the western Atlantic Ocean, whereas ACE-2 studied aerosols carried over the eastern Atlantic from both European urban/industrial and African mineral sources. These aerosols often have a marked influence on the top-of-atmosphere radiances measured by satellites. Shown there are contours of aerosol optical depth derived from radiances measured by the AVHRR sensor on the NOAA-11 satellite. The contours readily show that aerosols originating in North America, Europe, and Africa impact the radiative properties of air over the North Atlantic. However, the accurate derivation of flux changes, or radiative forcing, from the satellite measured radiances or retrieved optical depths remains a difficult challenge. In this paper we summarize key initial results from TARFOX and, to a lesser extent, ACE-2, with a focus on those results that allow an improved assessment of the flux changes caused by North Atlantic aerosols at middle latitudes.

  19. The S proteins of human coronavirus NL63 and severe acute respiratory syndrome coronavirus bind overlapping regions of ACE2.

    PubMed

    Li, Wenhui; Sui, Jianhua; Huang, I-Chueh; Kuhn, Jens H; Radoshitzky, Sheli R; Marasco, Wayne A; Choe, Hyeryun; Farzan, Michael

    2007-10-25

    The cellular receptor for human coronavirus NL63 (HCoV-NL63), a group I coronavirus, is angiotensin-converting enzyme2 (ACE2). ACE2 is also the receptor for the SARS coronavirus (SARS-CoV), a group II coronavirus. Here we describe the ability of HCoV-NL63 to utilize a number of ACE2 variants previously characterized as SARS-CoV receptors. Several ACE2 variants that reduced SARS-CoV S-protein association similarly reduced that of HCoV-NL63, whereas alteration of a number of solvent-exposed ACE2 residues did not interfere with binding by either S protein. One notable exception is ACE2 residue 354, at the boundary of the SARS-CoV binding site, whose alteration markedly inhibited utilization by the HCoV-NL63 but not SARS-CoV S proteins. In addition, the SARS-CoV S-protein receptor-binding domain inhibited entry mediated by the HCoV-NL63 S protein. These studies indicate that HCoV-NL63, like SARS-CoV, associates region of human ACE2 that includes a key loop formed by beta-strands 4 and 5.

  20. Wild-type and IL10-null mice have differential colonic epithelial gene expression responses to dietary supplementation with synbiotic Bifidobacterium animalis subspecies lactis and inulin.

    PubMed

    Kuo, Shiu-Ming; Chan, Wan-Chun; Hu, Zihua

    2014-03-01

    Prebiotic plus probiotic (synbiotic) supplementations promote fermentation and have shown anti-inflammatory activity in colonic epithelium. However, in many instances, patients with inflammatory bowel disease (IBD) have demonstrated adverse effects after prebiotic supplementation at a dose well tolerated by normal individuals. To test the hypothesis that the host inflammation affects the colonic epithelial response to increased fermentation, the gene expression of colonic epithelium was analyzed. In a 1-way experimental design to test the effect of supplements in wild-type mice using the standard diet formulated by the American Institute of Nutrition (AIN-93G) as the control diet, fermentable fiber inulin (5%) in the absence or presence of the probiotic Bifidobacterium animalis subspecies lactis (Bb12) (10(8) CFU/kg diet) showed limited effects on gene expression as determined by whole-genome microarray. Bb12 supplementation alone was known not to increase fermentation and here instead significantly upregulated genes in nucleic acid metabolic processes. The effects of the synbiotic diet were then determined in mice exposed to LPS-induced inflammation in a 2-way experimental design testing the effect of diet and LPS. The microarray and quantitative reverse transcription-polymerase chain reaction analyses on the wild-type mice revealed that LPS-induced changes in the colonic epithelium were 4- to 10-fold less in the synbiotic diet group compared with the control diet group. Unlike the wild-type mice, anti-inflammatory cytokine interleukin 10 (IL10)-null mice (susceptible to IBD) given the synbiotic diet, compared with those given the control diet, had 3- to 40-fold increased expression of inflammation-related genes such as Cxcl1 (chemokine C-X-C motif ligand 1) and S100a9 (S100 calcium binding protein A9) in the absence and presence of LPS exposure. These contrasting intestinal epithelial responses to increased fermentation in wild-type and IL10-null mice are similar

  1. Accumulation of xenotransplanted canine bone marrow cells in NOD/SCID/γc(null) mice with acute hepatitis induced by CCl4.

    PubMed

    Kato, Takashi; Hisasue, Masaharu; Segawa, Kazuhito; Fujimoto, Ayumi; Makiishi, Eri; Neo, Sakurako; Yasuno, Kyohei; Kobayashi, Ryosuke; Tsuchiya, Ryo

    2013-07-31

    Bone marrow cell infusion (BMI) has recently been suggested as an effective therapy for refractory liver disease; however, the efficiency of BMI using canine bone marrow cells (cBMCs) has not been reported. We evaluated the accumulation potential of cBMCs in a mouse model of acute liver failure. Acute hepatitis was induced by carbon tetrachloride (CCl4) treatment in NOD/SCID/γc(null)(NOG) mice and wild-type (WT) C57BL mice, and the characteristics of liver dysfunction and the degree of hepatic injury and regeneration were compared between the two mouse models. Next, female CCl4-treated NOG mice were xenotransplanted with male PKH26-labeled cBMCs, and the potential of cBMCs to accumulate in injured liver tissue compartments was examined. Fluorescence microscopy was performed to histologically detect the infused cBMCs, and DNA polymerase chain reaction was performed for detection of the male Y chromosome (SRY gene) in the recipient female NOG mice. The number of PKH26-positive cBMCs transplanted in the liver tissue gradually increased in the NOG mice. The infused cBMCs were located in the necrotic area of the liver at an early stage after transplantation, and most had accumulated a week after transplantation. However, the therapeutic efficacy of the xenotransplantation remained unclear, because no significant differences were observed concerning the extent liver injury and regeneration between the cBMC-transplanted and saline control mice. These results suggest that cBMCs will specifically accumulate in injured liver tissue and that BMC transplantation may have the potential to repair liver deficiency.

  2. A Single Nucleotide Polymorphism Uncovers a Novel Function for the Transcription Factor Ace2 during Candida albicans Hyphal Development

    PubMed Central

    Orellana-Muñoz, Sara; Gutiérrez-Escribano, Pilar; Arnáiz-Pita, Yolanda; Dueñas-Santero, Encarnación; Suárez, M. Belén; Bougnoux, Marie-Elisabeth; del Rey, Francisco; Sherlock, Gavin; d’Enfert, Christophe; Correa-Bordes, Jaime; de Aldana, Carlos R. Vázquez

    2015-01-01

    Candida albicans is a major invasive fungal pathogen in humans. An important virulence factor is its ability to switch between the yeast and hyphal forms, and these filamentous forms are important in tissue penetration and invasion. A common feature for filamentous growth is the ability to inhibit cell separation after cytokinesis, although it is poorly understood how this process is regulated developmentally. In C. albicans, the formation of filaments during hyphal growth requires changes in septin ring dynamics. In this work, we studied the functional relationship between septins and the transcription factor Ace2, which controls the expression of enzymes that catalyze septum degradation. We found that alternative translation initiation produces two Ace2 isoforms. While full-length Ace2, Ace2L, influences septin dynamics in a transcription-independent manner in hyphal cells but not in yeast cells, the use of methionine-55 as the initiation codon gives rise to Ace2S, which functions as the nuclear transcription factor required for the expression of cell separation genes. Genetic evidence indicates that Ace2L influences the incorporation of the Sep7 septin to hyphal septin rings in order to avoid inappropriate activation of cell separation during filamentous growth. Interestingly, a natural single nucleotide polymorphism (SNP) present in the C. albicans WO-1 background and other C. albicans commensal and clinical isolates generates a stop codon in the ninth codon of Ace2L that mimics the phenotype of cells lacking Ace2L. Finally, we report that Ace2L and Ace2S interact with the NDR kinase Cbk1 and that impairing activity of this kinase results in a defect in septin dynamics similar to that of hyphal cells lacking Ace2L. Together, our findings identify Ace2L and the NDR kinase Cbk1 as new elements of the signaling system that modify septin ring dynamics in hyphae to allow cell-chain formation, a feature that appears to have evolved in specific C. albicans lineages

  3. Null mutation of the β2 nicotinic acetylcholine receptor subunit attenuates nicotine withdrawal-induced anhedonia in mice.

    PubMed

    Stoker, Astrid K; Marks, Michael J; Markou, Athina

    2015-04-15

    The anhedonic signs of nicotine withdrawal are predictive of smoking relapse rates in humans. Identification of the neurobiological substrates that mediate anhedonia will provide insights into the genetic variations that underlie individual responses to smoking cessation and relapse. The present study assessed the role of β2 nicotinic acetylcholine receptor (nACh receptor) subunits in nicotine withdrawal-induced anhedonia using β2 nACh receptor subunit knockout (β2(-/-)) and wildtype (β2(+/+)) mice. Anhedonia was assessed with brain reward thresholds, defined as the current intensity that supports operant behavior in the discrete-trial current-intensity intracranial self-stimulation procedure. Nicotine was delivered chronically through osmotic minipumps for 28 days (40 mg/kg/day, base), and withdrawal was induced by either administering the broad-spectrum nicotinic receptor antagonist mecamylamine (i.e., antagonist-precipitated withdrawal) in mice chronically treated with nicotine or terminating chronic nicotine administration (i.e., spontaneous withdrawal). Mecamylamine (6 mg/kg, salt) significantly elevated brain reward thresholds in nicotine-treated β2(+/+) mice compared with saline-treated β2(+/+) mice and nicotine-treated β2(-/-) mice. Spontaneous nicotine withdrawal similarly resulted in significant elevations in thresholds in nicotine-withdrawing β2(+/+) mice compared with saline-treated β2(+/+) and nicotine-treated β2(-/-) mice, which remained at baseline levels. These results showed that precipitated and spontaneous nicotine withdrawal-induced anhedonia was attenuated in β2(-/-) mice. The reduced expression of anhedonic signs during nicotine withdrawal in β2(-/-) mice may have resulted from the lack of neuroadaptations in β2 nACh receptor subunit expression and function that may have occurred during either nicotine exposure or nicotine withdrawal in wildtype mice. In conclusion, individuals with genetic variations that result in diminished

  4. Maintenance of muscle mass and load-induced growth in Muscle RING Finger 1 null mice with age.

    PubMed

    Hwee, Darren T; Baehr, Leslie M; Philp, Andrew; Baar, Keith; Bodine, Sue C

    2014-02-01

    Age-related loss of muscle mass occurs to varying degrees in all individuals and has a detrimental effect on morbidity and mortality. Muscle RING Finger 1 (MuRF1), a muscle-specific E3 ubiquitin ligase, is believed to mediate muscle atrophy through the ubiquitin proteasome system (UPS). Deletion of MuRF1 (KO) in mice attenuates the loss of muscle mass following denervation, disuse, and glucocorticoid treatment; however, its role in age-related muscle loss is unknown. In this study, skeletal muscle from male wild-type (WT) and MuRF1 KO mice was studied up to the age of 24 months. Muscle mass and fiber cross-sectional area decreased significantly with age in WT, but not in KO mice. In aged WT muscle, significant decreases in proteasome activities, especially 20S and 26S β5 (20-40% decrease), were measured and were associated with significant increases in the maladaptive endoplasmic reticulum (ER) stress marker, CHOP. Conversely, in aged MuRF1 KO mice, 20S or 26S β5 proteasome activity was maintained or decreased to a lesser extent than in WT mice, and no increase in CHOP expression was measured. Examination of the growth response of older (18 months) mice to functional overload revealed that old WT mice had significantly less growth relative to young mice (1.37- vs. 1.83-fold), whereas old MuRF1 KO mice had a normal growth response (1.74- vs. 1.90-fold). These data collectively suggest that with age, MuRF1 plays an important role in the control of skeletal muscle mass and growth capacity through the regulation of cellular stress.

  5. Podocyte-Specific VEGF-A Gain of Function Induces Nodular Glomerulosclerosis in eNOS Null Mice

    PubMed Central

    Veron, Delma; Aggarwal, Pardeep K.; Velazquez, Heino; Kashgarian, Michael; Moeckel, Gilbert

    2014-01-01

    VEGF-A and nitric oxide are essential for glomerular filtration barrier homeostasis and are dysregulated in diabetic nephropathy. Here, we examined the effect of excess podocyte VEGF-A on the renal phenotype of endothelial nitric oxide synthase (eNOS) knockout mice. Podocyte-specific VEGF164 gain of function in eNOS−/− mice resulted in nodular glomerulosclerosis, mesangiolysis, microaneurysms, and arteriolar hyalinosis associated with massive proteinuria and renal failure in the absence of diabetic milieu or hypertension. In contrast, podocyte-specific VEGF164 gain of function in wild-type mice resulted in less pronounced albuminuria and increased creatinine clearance. Transmission electron microscopy revealed glomerular basement membrane thickening and podocyte effacement in eNOS−/− mice with podocyte-specific VEGF164 gain of function. Furthermore, glomerular nodules overexpressed collagen IV and laminin extensively. Biotin-switch and proximity ligation assays demonstrated that podocyte-specific VEGF164 gain of function decreased glomerular S-nitrosylation of laminin in eNOS−/− mice. In addition, treatment with VEGF-A decreased S-nitrosylated laminin in cultured podocytes. Collectively, these data indicate that excess glomerular VEGF-A and eNOS deficiency is necessary and sufficient to induce Kimmelstiel-Wilson–like nodular glomerulosclerosis in mice through a process that involves deposition of laminin and collagen IV and de-nitrosylation of laminin. PMID:24578128

  6. Behavioral characterization of cereblon forebrain-specific conditional null mice: a model for human non-syndromic intellectual disability.

    PubMed

    Rajadhyaksha, Anjali M; Ra, Stephen; Kishinevsky, Sarah; Lee, Anni S; Romanienko, Peter; DuBoff, Mariel; Yang, Chingwen; Zupan, Bojana; Byrne, Maureen; Daruwalla, Zeeba R; Mark, Willie; Kosofsky, Barry E; Toth, Miklos; Higgins, Joseph J

    2012-01-15

    A nonsense mutation in the human cereblon gene (CRBN) causes a mild type of autosomal recessive non-syndromic intellectual disability (ID). Animal studies show that crbn is a cytosolic protein with abundant expression in the hippocampus (HPC) and neocortex (CTX). Its diverse functions include the developmental regulation of ion channels at the neuronal synapse, the mediation of developmental programs by ubiquitination, and a target for herpes simplex type I virus in HPC neurons. To test the hypothesis that anomalous CRBN expression leads to HPC-mediated memory and learning deficits, we generated germ-line crbn knock-out mice (crbn(-/-)). We also inactivated crbn in forebrain neurons in conditional knock-out mice in which crbn exons 3 and 4 are deleted by cre recombinase under the direction of the Ca(2+)/calmodulin-dependent protein kinase II alpha promoter (CamKII(cre/+), crbn(-/-)). crbn mRNA levels were negligible in the HPC, CTX, and cerebellum (CRBM) of the crbn(-/-) mice. In contrast, crbn mRNA levels were reduced 3- to 4-fold in the HPC, CTX but not in the CRBM in CamKII(cre/+), crbn(-/-) mice as compared to wild type (CamKII(cre/+), crbn(+/+)). Contextual fear conditioning showed a significant decrease in the percentage of freezing time in CamKII(cre/+), crbn(-/-) and crbn(-/-) mice while motor function, exploratory motivation, and anxiety-related behaviors were normal. These findings suggest that CamKII(cre/+), crbn(-/-) mice exhibit selective HPC-dependent deficits in associative learning and supports the use of these mice as in vivo models to study the functional consequences of CRBN aberrations on memory and learning in humans.

  7. Determining the Enzymatic Activity of Angiotensin-Converting Enzyme 2 (ACE2) in Brain Tissue and Cerebrospinal Fluid Using a Quenched Fluorescent Substrate.

    PubMed

    Sriramula, Srinivas; Pedersen, Kim Brint; Xia, Huijing; Lazartigues, Eric

    2017-01-01

    Angiotensin-converting enzyme 2 (ACE2) is a component of the renin-angiotensin system (RAS) which plays an important role in the regulation of blood pressure and volume homeostasis. Accumulating evidence shows alterations in ACE2 expression and activity in several hypertensive animal models, as well as in patients with hypertension. In order to assess the role of brain ACE2 in hypertension, a specific ACE2 assay is required. Based on a quenched fluorescent substrate, we describe an easy-to-use method for determining ACE2 activity in brain tissue and cerebrospinal fluid. The method can further be adapted for other tissues, plasma, cell extracts, and cell culture supernatants.

  8. Sensorineural hearing loss in insulin-like growth factor I-null mice: a new model of human deafness.

    PubMed

    Cediel, R; Riquelme, R; Contreras, J; Díaz, A; Varela-Nieto, I

    2006-01-01

    It has been reported that mutations in the gene encoding human insulin-like growth factor-I (IGF-I) cause syndromic hearing loss. To study the precise role of IGF-I in auditory function and to hypothesize the possible morphological and electrophysiological changes that may occur in the human inner ear, we have analysed the auditory brainstem response in a mouse model of IGF-I deficiency. We show here that homozygous Igf-1(-/-) mice present an all-frequency involved bilateral sensorineural hearing loss. Igf-1(-/-) mice also present a delayed response to acoustic stimuli; this increases along the auditory pathway, indicating a contribution of the central nervous system to the hearing loss in Igf-1(-/-) mice. These results support the use of the Igf-1(-/-) mouse as a new model for the study of human syndromic deafness.

  9. ACE2, angiotensin-(1-7) and Mas receptor axis in inflammation and fibrosis

    PubMed Central

    Simões e Silva, AC; Silveira, KD; Ferreira, AJ; Teixeira, MM

    2013-01-01

    Recent advances have improved our understanding of the renin-angiotensin system (RAS). These have included the recognition that angiotensin (Ang)-(1-7) is a biologically active product of the RAS cascade. The identification of the ACE homologue ACE2, which forms Ang-(1-7) from Ang II, and the GPCR Mas as an Ang-(1-7) receptor have provided the necessary biochemical and molecular background and tools to study the biological significance of Ang-(1-7). Most available evidence supports a counter-regulatory role for Ang-(1-7) by opposing many actions of Ang II on AT1 receptors, especially vasoconstriction and proliferation. Many studies have now shown that Ang-(1-7) by acting via Mas receptor exerts inhibitory effects on inflammation and on vascular and cellular growth mechanisms. Ang-(1-7) has also been shown to reduce key signalling pathways and molecules thought to be relevant for fibrogenesis. Here, we review recent findings related to the function of the ACE2/Ang-(1-7)/Mas axis and focus on the role of this axis in modifying processes associated with acute and chronic inflammation, including leukocyte influx, fibrogenesis and proliferation of certain cell types. More attention will be given to the involvement of the ACE2/Ang-(1-7)/Mas axis in the context of renal disease because of the known relevance of the RAS for the function of this organ and for the regulation of kidney inflammation and fibrosis. Taken together, this knowledge may help in paving the way for the development of novel treatments for chronic inflammatory and renal diseases. PMID:23488800

  10. Desert Dust Layers Over Polluted Marine Boundary Layers: ACE-2 Measurements and ACE-Asia Plans

    NASA Technical Reports Server (NTRS)

    Russell, Philip B.; Schmid, B.; Livingston, J. M.; Redemann, J.; Bergstrom, R. W.; Condon, Estelle P. (Technical Monitor)

    2000-01-01

    Aerosols in ACE-Asia are expected to have some commonalties with those in ACE-2, along with important differences. Among the commonalities are occurrences of desert dust layers over polluted marine boundary layers. Differences include the nature of the dust (yellowish in the East Asia desert outflow, vs. reddish-brown in the Sahara Outflow measured in ACE-2) and the composition of boundary-layer aerosols (e.g., more absorbing, soot and organic aerosol in-the Asian plume, caused by coal and biomass burning, with limited controls). In this paper we present ACE-2 measurements and analyses as a guide to our plans for ACE-2 Asia. The measurements include: (1) Vertical profiles of aerosol optical depth and extinction (380-1558 nm), and of water vapor column and concentration, from the surface through the elevated desert dust, measured by the 14-channel Ames Airborne Tracking Sunphotometer (AATS-14); (2) Comparisons of airborne and shipborne sunphotometer optical depths to satellite-retrieved values, with and without desert dust; (3) Comparisons between airborne Sunphotometer optical depth and extinction spectra and those derived from coincident airborne in situ measurements of aerosol size distribution, scattering and absorption; (4) Comparisons between size distributions measured in situ and retrieved from sunphotometer optical depth spectra; (5) Comparisons between aerosol single scattering albedo values obtained by several techniques, using various combinations of measurements of backscatter, extinction, size distribution, scattering, absorption, and radiative flux. We show how analyses of these data can be used to address questions important to ACE-Asia, such as: (1) How do dust and other absorbing aerosols affect the accuracy of satellite optical depth retrievals? How important are asphericity effects? (2) How important are supermicron dust and seasalt aerosols to overall aerosol optical depth and radiative forcing? How well are these aerosols sampled by aircraft

  11. Assessment of functional recovery and axonal sprouting in oligodendrocyte-myelin glycoprotein (OMgp) null mice after spinal cord injury.

    PubMed

    Ji, Benxiu; Case, Lauren C; Liu, Kai; Shao, Zhaohui; Lee, Xinhua; Yang, Zhongshu; Wang, Joy; Tian, Tim; Shulga-Morskaya, Svetlana; Scott, Martin; He, Zhigang; Relton, Jane K; Mi, Sha

    2008-10-01

    Oligodendrocyte-myelin glycoprotein (OMgp) is a myelin component that has been shown in vitro to inhibit neurite outgrowth by binding to the Nogo-66 receptor (NgR1)/Lingo-1/Taj (TROY)/p75 receptor complex to activate the RhoA pathway. To investigate the effects of OMgp on axon regeneration in vivo, OMgp(-/-) mice on a mixed 129/Sv/C57BL/6 (129BL6) or a C57BL/6 (BL6) genetic background were tested in two spinal cord injury (SCI) models - a severe complete transection or a milder dorsal hemisection. OMgp(-/-) mice on the mixed 129BL6 genetic background showed greater functional improvement compared to OMgp(+/+) littermates, with increased numbers of cholera toxin B-labeled ascending sensory axons and 5-HT(+) descending axons and less RhoA activation after spinal cord injury. Myelin isolated from OMgp(-/-) mice (129BL6) was significantly less inhibitory to neurite outgrowth than wild-type (wt) myelin in vitro. However, OMgp(-/-) mice on a BL/6 genetic background showed neither statistically significant functional recovery nor axonal sprouting following dorsal hemisection.

  12. Glutamate Cysteine Ligase Modifier Subunit (Gclm) Null Mice Have Increased Ovarian Oxidative Stress and Accelerated Age-Related Ovarian Failure

    PubMed Central

    Lim, Jinhwan; Nakamura, Brooke N.; Mohar, Isaac; Kavanagh, Terrance J.

    2015-01-01

    Glutathione (GSH) is the one of the most abundant intracellular antioxidants. Mice lacking the modifier subunit of glutamate cysteine ligase (Gclm), the rate-limiting enzyme in GSH synthesis, have decreased GSH. Our prior work showed that GSH plays antiapoptotic roles in ovarian follicles. We hypothesized that Gclm−/− mice have accelerated ovarian aging due to ovarian oxidative stress. We found significantly decreased ovarian GSH concentrations and oxidized GSH/oxidized glutathione redox potential in Gclm−/− vs Gclm+/+ ovaries. Prepubertal Gclm−/− and Gclm+/+ mice had similar numbers of ovarian follicles, and as expected, the total number of ovarian follicles declined with age in both genotypes. However, the rate of decline in follicles was significantly more rapid in Gclm−/− mice, and this was driven by accelerated declines in primordial follicles, which constitute the ovarian reserve. We found significantly increased 4-hydroxynonenal immunostaining (oxidative lipid damage marker) and significantly increased nitrotyrosine immunostaining (oxidative protein damage marker) in prepubertal and adult Gclm−/− ovaries compared with controls. The percentage of small ovarian follicles with increased granulosa cell proliferation was significantly higher in prepubertal and 2-month-old Gclm−/− vs Gclm+/+ ovaries, indicating accelerated recruitment of primordial follicles into the growing pool. The percentages of growing follicles with apoptotic granulosa cells were increased in young adult ovaries. Our results demonstrate increased ovarian oxidative stress and oxidative damage in young Gclm−/− mice, associated with an accelerated decline in ovarian follicles that appears to be mediated by increased recruitment of follicles into the growing pool, followed by apoptosis at later stages of follicular development. PMID:26083875

  13. Disturbances in the secretion of neurotransmitters in IA-2/IA-2beta null mice: changes in behavior, learning and lifespan.

    PubMed

    Nishimura, T; Kubosaki, A; Ito, Y; Notkins, A L

    2009-03-17

    Islet-associated protein 2 (IA-2) and IA-2beta are major autoantigens in type 1 diabetes and transmembrane proteins in dense core secretory vesicles (DCV) of neuroendocrine cells. The deletion of these genes results in a decrease in insulin secretion. The present study was initiated to test the hypothesis that this deletion not only affects the secretion of insulin, but has a more global effect on neuroendocrine secretion that leads to disturbances in behavior and learning. Measurement of neurotransmitters showed that norepinephrine, dopamine and 5-HT were significantly decreased in the brain of double knockout (DKO) mice (P<0.05 to <0.001). In tests evaluating anxiety-like behavior and conditioned-learning, the DKO mice showed a highly significant increase in anxiety-like behavior (P<0.01 to <0.001) and impairment of conditioned learning (P<0.01) as compared to WT mice. The DKO mice also displayed an increase in spontaneous and induced seizures (P<0.01) and age-related death. Contrary to the generally held view that IA-2 and IA-2beta are expressed exclusively in DCV, subcellular fractionation studies revealed that IA-2beta, but not IA-2, co-purifies with fractions rich in synaptic vesicles (SV), and that the secretion of dopamine, GABA and glutamate from the synaptosomes of the DKO mice was significantly decreased as was the number of SV (P<0.01). Taken together, these findings show that IA-2beta is present in both DCV and SV, and that the deletion of IA-2/IA-2beta has a global effect on the secretion of neurotransmitters. The impairment of secretion leads to behavioral and learning disturbances, seizures and reduced lifespan.

  14. Presenilin-1 familial Alzheimer's disease mutation alters hippocampal neurogenesis and memory function in CCL2 null mice.

    PubMed

    Kiyota, Tomomi; Morrison, Christine M; Tu, Guihua; Dyavarshetty, Bhagyalaxmi; Weir, Robert A; Zhang, Gang; Xiong, Huangui; Gendelman, Howard E

    2015-10-01

    Aberrations in hippocampal neurogenesis are associated with learning and memory, synaptic plasticity and neurodegeneration in Alzheimer's disease (AD). However, the linkage between them, β-amyloidosis and neuroinflammation is not well understood. To this end, we generated a mouse overexpressing familial AD (FAD) mutant human presenilin-1 (PS1) crossed with a knockout (KO) of the CC-chemokine ligand 2 (CCL2) gene. The PS1/CCL2KO mice developed robust age-dependent deficits in hippocampal neurogenesis associated with impairments in learning and memory, synaptic plasticity and long-term potentiation. Neurogliogenesis gene profiling supported β-amyloid independent pathways for FAD-associated deficits in hippocampal neurogenesis. We conclude that these PS1/CCL2KO mice are suitable for studies linking host genetics, immunity and hippocampal function.

  15. Autistic-like syndrome in mu opioid receptor null mice is relieved by facilitated mGluR4 activity.

    PubMed

    Becker, Jérôme A J; Clesse, Daniel; Spiegelhalter, Coralie; Schwab, Yannick; Le Merrer, Julie; Kieffer, Brigitte L

    2014-08-01

    The etiology of Autism Spectrum Disorders (ASDs) remains largely unknown. Identifying vulnerability genes for autism represents a major challenge in the field and allows the development of animal models for translational research. Mice lacking the mu opioid receptor gene (Oprm1(-/-)) were recently proposed as a monogenic mouse model of autism, based on severe deficits in social behavior and communication skills. We confirm this hypothesis by showing that adult Oprm1(-/-) animals recapitulate core and multiple comorbid behavioral symptoms of autism and also display anatomical, neurochemical, and genetic landmarks of the disease. Chronic facilitation of mGluR4 signaling, which we identified as a novel pharmacological target in ASDs in these mice, was more efficient in alleviating behavioral deficits than the reference molecule risperidone. Altogether, our data provide first evidence that disrupted mu opioid receptor signaling is sufficient to trigger a comprehensive autistic syndrome, maybe through blunted social reward processes, and this mouse model opens promising avenues for therapeutic innovation.

  16. Mouse and human BAC transgenes recapitulate tissue-specific expression of the vitamin D receptor in mice and rescue the VDR-null phenotype.

    PubMed

    Lee, Seong Min; Bishop, Kathleen A; Goellner, Joseph J; O'Brien, Charles A; Pike, J Wesley

    2014-06-01

    The biological actions of 1,25-dihydroxyvitamin D3 (1,25(OH)2D3) are mediated by the vitamin D receptor (VDR), which is expressed in numerous target tissues in a cell type-selective manner. Recent studies using genomic analyses and recombineered bacterial artificial chromosomes (BACs) have defined the specific features of mouse and human VDR gene loci in vitro. In the current study, we introduced recombineered mouse and human VDR BACs as transgenes into mice and explored their expression capabilities in vivo. Individual transgenic mouse strains selectively expressed BAC-derived mouse or human VDR proteins in appropriate vitamin D target tissues, thereby recapitulating the tissue-specific expression of endogenous mouse VDR. The mouse VDR transgene was also regulated by 1,25(OH)2D3 and dibutyryl-cAMP. When crossed into a VDR-null mouse background, both transgenes restored wild-type basal as well as 1,25(OH)2D3-inducible gene expression patterns in the appropriate tissues. This maneuver resulted in the complete rescue of the aberrant phenotype noted in the VDR-null mouse, including systemic features associated with altered calcium and phosphorus homeostasis and disrupted production of parathyroid hormone and fibroblast growth factor 23, and abnormalities associated with the skeleton, kidney, parathyroid gland, and the skin. This study suggests that both mouse and human VDR transgenes are capable of recapitulating basal and regulated expression of the VDR in the appropriate mouse tissues and restore 1,25(OH)2D3 function. These results provide a baseline for further dissection of mechanisms integral to mouse and human VDR gene expression and offer the potential to explore the consequence of selective mutations in VDR proteins in vivo.

  17. Abnormality in Wnt Signaling is Causatively Associated with Oxidative Stress-Induced Intestinal Tumorigenesis in MUTYH-Null Mice

    PubMed Central

    Isoda, Takuro; Nakatsu, Yoshimichi; Yamauchi, Kazumi; Piao, Jingshu; Yao, Takashi; Honda, Hiroshi; Nakabeppu, Yusaku; Tsuzuki, Teruhisa

    2014-01-01

    MUTYH is a DNA glycosylase that excises adenine paired with 8-oxoguanine to prevent mutagenesis in mammals. Biallelic germline mutations of MUTYH have been found in patients predisposed to a recessive form of familial adenomatous polyposis (MAP: MUTYH-associated polyposis). We previously reported that Mutyh-deficient mice showed a high susceptibility to spontaneous and oxidative stress-induced intestinal adenoma/carcinoma. Here, we performed mutation analysis of the tumor-associated genes including Apc, Ctnnb1, Kras and Trp53 in the intestinal tumors of Mutyh-deficient mice. In the 62 tumors, we identified 25 mutations in Apc of 18 tumors and 36 mutations in Ctnnb1 of 36 tumors. Altogether, 54 out of the 62 tumors (87.1%) had a mutation in either Apc or Ctnnb1; no tumor displayed mutations simultaneously in the both genes. Similar to MAP, 60 out of 61 mutations (98.3%) were identified as G:C to T:A transversions of which 85% occurred at either AGAA or TGAA sequences. Immunohistochemical analyses revealed the accumulation of β-catenin in the nuclei of tumors. No mutation was found in either Kras or Trp53 in the tumors. These results indicate that the uncontrolled activation of Wnt signaling pathway is causatively associated with oxidative stress-induced intestinal tumorigenesis in the Mutyh-deficient mice. PMID:25170306

  18. Distinct Luminal-Type Mammary Carcinomas Arise from Orthotopic Trp53-Null Mammary Transplantation of Juvenile versus Adult Mice

    DOE PAGES

    Nguyen, David H.; Ouyang, Haoxu; Mao, Jian-Hua; ...

    2014-12-01

    Age and physiologic status, such as menopause, are risk factors for breast cancer. Less clear is what factors influence the diversity of breast cancer. In this study, we investigated the effect of host age on the distribution of tumor subtypes in mouse mammary chimera consisting of wild-type hosts and Trp53 nullizygous epithelium, which undergoes a high rate of neoplastic transformation. Wild-type mammary glands cleared of endogenous epithelium at 3 weeks of age were subsequently transplanted during puberty (5 weeks) or at maturation (10 weeks) with syngeneic Trp53-null mammary tissue fragments and monitored for one year. Tumors arose sooner from adultmore » hosts (AH) compared with juvenile hosts (JH). However, compared with AH tumors, JH tumors grew several times faster, were more perfused, exhibited a two-fold higher mitotic index, and were more highly positive for insulin-like growth factor receptor phosphorylation. Most tumors in each setting were estrogen receptor (ER)-positive (80% JH vs. 70% AH), but JH tumors were significantly more ER-immunoreactive (P = 0.0001) than AH tumors. A differential expression signature (JvA) of juvenile versus adult tumors revealed a luminal transcriptional program. Centroids of the human homologs of JvA genes showed that JH tumors were more like luminal A tumors and AH tumors were more like luminal B tumors. Hierarchical clustering with the JvA human ortholog gene list segregated luminal A and luminal B breast cancers across datasets. Lastly, these data support the notion that age-associated host physiology greatly influences the intrinsic subtype of breast cancer.« less

  19. Distinct Luminal-Type Mammary Carcinomas Arise from Orthotopic Trp53-Null Mammary Transplantation of Juvenile versus Adult Mice

    SciTech Connect

    Nguyen, David H.; Ouyang, Haoxu; Mao, Jian-Hua; Hlatky, Lynn; Barcellos-Hoff, M. H.

    2014-12-01

    Age and physiologic status, such as menopause, are risk factors for breast cancer. Less clear is what factors influence the diversity of breast cancer. In this study, we investigated the effect of host age on the distribution of tumor subtypes in mouse mammary chimera consisting of wild-type hosts and Trp53 nullizygous epithelium, which undergoes a high rate of neoplastic transformation. Wild-type mammary glands cleared of endogenous epithelium at 3 weeks of age were subsequently transplanted during puberty (5 weeks) or at maturation (10 weeks) with syngeneic Trp53-null mammary tissue fragments and monitored for one year. Tumors arose sooner from adult hosts (AH) compared with juvenile hosts (JH). However, compared with AH tumors, JH tumors grew several times faster, were more perfused, exhibited a two-fold higher mitotic index, and were more highly positive for insulin-like growth factor receptor phosphorylation. Most tumors in each setting were estrogen receptor (ER)-positive (80% JH vs. 70% AH), but JH tumors were significantly more ER-immunoreactive (P = 0.0001) than AH tumors. A differential expression signature (JvA) of juvenile versus adult tumors revealed a luminal transcriptional program. Centroids of the human homologs of JvA genes showed that JH tumors were more like luminal A tumors and AH tumors were more like luminal B tumors. Hierarchical clustering with the JvA human ortholog gene list segregated luminal A and luminal B breast cancers across datasets. Lastly, these data support the notion that age-associated host physiology greatly influences the intrinsic subtype of breast cancer.

  20. Beneficial effects of quinoline-3-carboxamide (ABR-215757) on atherosclerotic plaque morphology in S100A12 transgenic ApoE null mice

    PubMed Central

    Yan, Ling; Bjork, Per; Butuc, Radu; Gawdzik, Joseph; Earley, Judy; Kim, Gene; Hofmann Bowman, Marion A

    2013-01-01

    Objective There is an emerging widespread interest in the role of damage-associated molecular pattern molecules (DAMP) S100A8, S100A9 and S100A12 in cardiovascular and other diseases. In this study we tested the efficacy of ABR-215757, a S100 protein binding immuno-modulatory compound to stabilize atherosclerosis in transgenic ApoE null mice that express the human pro-inflammatory S100A12 protein within the smooth muscle cell (SM22α-S100A12). Methods Twelve-week old S100A12 transgenic/ApoE-/- and WT/ApoE-/- mice were treated with ABR-21575 for 5 weeks and were analyzed 4 month later. Results Surface plasmon resonance analysis demonstrated that S100A12 interacts with ABR-215757 in a zinc dependent manner in vitro. In vivo, ABR-215757 administration reduced features of advanced plaque morphology resulting in smaller necrotic cores, diminished intimal and medial vascular calcification, and reduced amount of infiltrating inflammatory cells. ABR-215757 normalized aortic expression of RAGE protein and normalized experimentally-induced delayed hypersensitivity. The effect of ABR-215757 was more prominent in ApoE-/- mice expressing S100A12 than in ApoE-/- animals lacking expression of human S100A12 protein. Conclusion Our data suggest that S100A12 is important for progression of atherosclerosis and can be targeted by the small molecule ABR-215757. The specific binding of quinoline-3-carboxamides to S100A12 attenuates S100A12-mediated features of accelerated murine atherosclerosis. PMID:23497784

  1. Highly efficient in vivo delivery of PMO into regenerating myotubes and rescue in laminin-α2 chain-null congenital muscular dystrophy mice.

    PubMed

    Aoki, Yoshitsugu; Nagata, Tetsuya; Yokota, Toshifumi; Nakamura, Akinori; Wood, Matthew J A; Partridge, Terence; Takeda, Shin'ichi

    2013-12-15

    Phosphorodiamidate morpholino oligomer (PMO)-mediated exon skipping is among the more promising approaches to the treatment of several neuromuscular disorders including Duchenne muscular dystrophy. The main weakness of this approach arises from the low efficiency and sporadic nature of the delivery of charge-neutral PMO into muscle fibers, the mechanism of which is unknown. In this study, to test our hypothesis that muscle fibers take up PMO more efficiently during myotube formation, we induced synchronous muscle regeneration by injection of cardiotoxin into the tibialis anterior muscle of Dmd exon 52-deficient mdx52 and wild-type mice. Interestingly, by in situ hybridization, we detected PMO mainly in embryonic myosin heavy chain-positive regenerating fibers. In addition, we showed that PMO or 2'-O-methyl phosphorothioate is taken up efficiently into C2C12 myotubes when transfected 24-72 h after the induction of differentiation but is poorly taken up into undifferentiated C2C12 myoblasts suggesting efficient uptake of PMO in the early stages of C2C12 myotube formation. Next, we tested the therapeutic potential of PMO for laminin-α2 chain-null dy(3K)/dy(3K) mice: a model of merosin-deficient congenital muscular dystrophy (MDC1A) with active muscle regeneration. We confirmed the recovery of laminin-α2 chain and slightly prolonged life span following skipping of the mutated exon 4 in dy(3K)/dy(3K) mice. These findings support the idea that PMO entry into fibers is dependent on a developmental stage in myogenesis rather than on dystrophinless muscle membranes and provide a platform for developing PMO-mediated therapies for a variety of muscular disorders, such as MDC1A, that involve active muscle regeneration.

  2. A comparative study of the central effects of specific POMC-derived melanocortin peptides on food intake and body weight in Pomc null mice

    PubMed Central

    Tung, YC Loraine; Piper, Sarah J; Yeung, Debra; O’Rahilly, Stephen; Coll, Anthony P

    2008-01-01

    Functional disruption of either MC3R or MC4R results in obesity, implicating both in the control of energy homeostasis. The ligands for these receptors are derived from the prohormone POMC, which is post-translationally processed to produce a set of melanocortin peptides with a range of activities at the MC3R and MC4R. The relative importance of each of these peptides (α-MSH, γ3 MSH, γ2 MSH, γ lipotropin (γ-LPH) and, in man but not in rodents, β-MSH) in the maintenance of energy homeostasis is, as yet, unclear. To investigate this further, equimolar amounts (2nmols) of each peptide were centrally administered to freely feeding, corticosterone supplemented, Pomc null (Pomc-/-) mice. Following a single dose at the onset of the dark cycle, α-MSH had the most potent anorexigenic effect, reducing food intake to 35% of sham treated animals. β-MSH, γ-LPH, γ3-and γ2- MSH all reduced food intake but to a lesser degree. The effects of peptide administration over three days were also assessed. Only α-MSH significantly reduced body weight, affecting both fat and lean mass. Other peptides had no significant effect on body weight. Pair-feeding of sham-treated mice to those treated with α-MSH resulted in identical changes in total weight, fat and lean mass indicating that the effects of α-MSH were primarily due to reduced food intake rather than increased energy expenditure. Although other melanocortins can reduce food intake in the short term, only α-MSH can reduce the excess fat and lean mass found in Pomc-/- mice, mediated largely through an effect on food intake. PMID:16959830

  3. Resilient RTN fast spiking in Kv3.1 null mice suggests redundancy in the action potential repolarization mechanism.

    PubMed

    Porcello, Darrell M; Ho, Chi Shun; Joho, Rolf H; Huguenard, John R

    2002-03-01

    Fast spiking (FS), GABAergic neurons of the reticular thalamic nucleus (RTN) are capable of firing high-frequency trains of brief action potentials, with little adaptation. Studies in recombinant systems have shown that high-voltage-activated K(+) channels containing the Kv3.1 and/or Kv3.2 subunits display biophysical properties that may contribute to the FS phenotype. Given that RTN expresses high levels of Kv3.1, with little or no Kv3.2, we tested whether this subunit was required for the fast action potential repolarization mechanism essential to the FS phenotype. Single- and multiple-action potentials were recorded using whole-cell current clamp in RTN neurons from brain slices of wild-type and Kv3.1-deficient mice. At 23 degrees C, action potentials recorded from homozygous Kv3.1 deficient mice (Kv3.1(-/-)) compared with their wild-type (Kv3.1(+/+)) counterparts had reduced amplitudes (-6%) and fast after-hyperpolarizations (-16%). At 34 degrees C, action potentials in Kv3.1(-/-) mice had increased duration (21%) due to a reduced rate of repolarization (-30%) when compared with wild-type controls. Action potential trains in Kv3.1(-/-) were associated with a significantly greater spike decrement and broadening and a diminished firing frequency versus injected current relationship (F/I) at 34 degrees C. There was no change in either spike count or maximum instantaneous frequency during low-threshold Ca(2+) bursts in Kv3.1(-/-) RTN neurons at either temperature tested. Our findings show that Kv3.1 is not solely responsible for fast spikes or high-frequency firing in RTN neurons. This suggests genetic redundancy in the system, possibly in the form of other Kv3 members, which may suffice to maintain the FS phenotype in RTN neurons in the absence of Kv3.1.

  4. Disruption of NBS1 gene leads to early embryonic lethality in homozygous null mice and induces specific cancer in heterozygous mice

    SciTech Connect

    Kurimasa, Akihiro; Burma, Sandeep; Henrie, Melinda; Ouyang, Honghai; Osaki, Mitsuhiko; Ito, Hisao; Nagasawa, Hatsumi; Little, John B.; Oshimura, Mitsuo; Li, Gloria C.; Chen, David J.

    2002-04-15

    Nijmegen breakage syndrome (NBS) is a rare autosomal recessive chromosome instability syndrome characterized by microcephaly, growth retardation, immunodeficiency, and cancer predisposition, with cellular features similar to that of ataxia telangiectasia (AT). NBS results from mutations in the mammalian gene Nbs1 that codes for a 95-kDa protein called nibrin, NBS1, or p95. To establish an animal model for NBS, we attempted to generate NBS1 knockout mice. However, NBS1 gene knockouts were lethal at an early embryonic stage. NBS1 homozygous(-/-) blastocyst cells cultured in vitro showed retarded growth and subsequently underwent growth arrest within 5 days of culture. Apoptosis, assayed by TUNEL staining, was observed in NBSI homozygous(-/-) blastocyst cells cultured for four days. NBSI heterozygous(+/-) mice were normal, and exhibited no specific phenotype for at least one year. However, fibroblast cells from NBSI heterozygous(+/-) mice displayed an enhanced frequency of spontaneous transformation to anchorage-independent growth as compared to NBS1 wild-type(+/+) cells. Furthermore, heterozygous(+/-) mice exhibited a high incidence of hepatocellular carcinoma after one year compared to wild-type mice, even though no significant differences in the incidence of other tumors such as lung adenocarcinoma and lymphoma were observed. Taken together, these results strongly suggest that NBS1 heterozygosity and reduced NBSI expression induces formation of specific tumors in mice.

  5. Mechanical and material properties of cortical and trabecular bone from cannabinoid receptor-1-null (Cnr1(-/-)) mice.

    PubMed

    Khalid, Aysha B; Goodyear, Simon R; Ross, Ruth A; Aspden, Richard M

    2016-10-01

    The endocannabinoid system is known for its regulatory effects on bone metabolism through the cannabinoid receptors, Cnr1 and Cnr2. In this study we analysed the mechanical and material properties of long bones from Cnr1(-/-) mice on a C57BL/6 background. Tibiae and femora from 5- and 12-week-old mice were subjected to three-point bending to measure bending stiffness and yield strength. Elastic modulus, density and mineral content were measured in the diaphysis. Second moment of area (MOA2), inner and outer perimeters of the cortical shaft and trabecular fractional bone volume (BV/TV) were measured using micro-CT. In Cnr1(-/-) males and females at both ages the bending stiffness was reduced due to a smaller MOA2. Bone from Cnr1(-/-) females had a greater modulus than wild-type controls, although no differences were observed in males. BV/TV of 12-week-old Cnr1(-/-) females was greater than controls, although no difference was seen at 5-weeks. On the contrary, Cnr1(-/-) males had the same BV/TV as controls at 12-weeks while they had significantly lower values at 5-weeks. This study shows that deleting Cnr1 decreases the amount of cortical bone in both males and females at 12-weeks, but increases the amount of trabecular bone only in females.

  6. Sex differences in brain proteomes of neuron-specific STAT3-null mice after cerebral ischemia/reperfusion.

    PubMed

    Di Domenico, Fabio; Casalena, Gabriella; Jia, Jia; Sultana, Rukhsana; Barone, Eugenio; Cai, Jian; Pierce, William M; Cini, Chiara; Mancuso, Cesare; Perluigi, Marzia; Davis, Catherine M; Alkayed, Nabil J; Butterfield, D Allan; Butterfield, Allan D

    2012-05-01

    Signal transduction and activator of transcription-3 (STAT3) plays an important role in neuronal survival, regeneration and repair after brain injury. We previously demonstrated that STAT3 is activated in brain after cerebral ischemia specifically in neurons. The effect was sex-specific and modulated by sex steroids, with higher activation in females than males. In the current study, we used a proteomics approach to identify downstream proteins affected by ischemia in male and female wild-type (WT) and neuron-specific STAT3 knockout (KO) mice. We established four comparison groups based on the transgenic condition and the hemisphere analyzed, respectively. Moreover, the sexual variable was taken into account and male and female animals were analyzed independently. Results support a role for STAT3 in metabolic, synaptic, structural and transcriptional responses to cerebral ischemia, indeed the adaptive response to ischemia/reperfusion injury is delayed in neuronal-specific STAT3 KO mice. The differences observed between males and females emphasize the importance of sex-specific neuronal survival and repair mechanisms, especially those involving antioxidant and energy-related activities, often caused by sex hormones.

  7. The transcription factor HNF1α induces expression of angiotensin-converting enzyme 2 (ACE2) in pancreatic islets from evolutionarily conserved promoter motifs.

    PubMed

    Pedersen, Kim Brint; Chhabra, Kavaljit H; Nguyen, Van K; Xia, Huijing; Lazartigues, Eric

    2013-11-01

    Pancreatic angiotensin-converting enzyme 2 (ACE2) has previously been shown to be critical for maintaining glycemia and β-cell function. Efforts to maintain or increase ACE2 expression in pancreatic β-cells might therefore have therapeutic potential for treating diabetes. In our study, we investigated the transcriptional role of hepatocyte nuclear factor 1α (HNF1α) and hepatocyte nuclear factor 1β (HNF1β) in induction of ACE2 expression in insulin-secreting cells. A deficient allele of HNF1α or HNF1β causes maturity-onset diabetes of the young (MODY) types 3 and 5, respectively, in humans. We found that ACE2 is primarily transcribed from the proximal part of the ACE2 promoter in the pancreas. In the proximal part of the human ACE2 promoter, we further identified three functional HNF1 binding sites, as they have binding affinity for HNF1α and HNF1β and are required for induction of promoter activity by HNF1β in insulinoma cells. These three sites are well-conserved among mammalian species. Both HNF1α and HNF1β induce expression of ACE2 mRNA and lead to elevated levels of ACE2 protein and ACE2 enzymatic activity in insulinoma cells. Furthermore, HNF1α dose-dependently increases ACE2 expression in primary pancreatic islet cells. We conclude that HNF1α can induce the expression of ACE2 in pancreatic islet cells via evolutionarily conserved HNF1 binding sites in the ACE2 promoter. Potential therapeutics aimed at counteracting functional HNF1α depletion in diabetes and MODY3 will thus have ACE2 induction in pancreatic islets as a likely beneficial effect.

  8. Polymorphisms of ACE2 gene are associated with essential hypertension and antihypertensive effects of Captopril in women.

    PubMed

    Fan, X; Wang, Y; Sun, K; Zhang, W; Yang, X; Wang, S; Zhen, Y; Wang, J; Li, W; Han, Y; Liu, T; Wang, X; Chen, J; Wu, H; Hui, R

    2007-08-01

    ACE2 appears to counterbalance the vasopressor effect of angiotensin I converting enzyme (ACE) in the reninangiotensin system. We hypothesized that ACE2 polymorphisms could confer a high risk of hypertension and have an impact on the antihypertensive response to ACE inhibitors. The hypothesis was tested in two casecontrol studies and a clinical trial of 3,408 untreated hypertensive patients randomized to Atenolol, Hydrochlorothiazide, Captopril, or Nifedipine treatments for 4 weeks. ACE2 rs2106809 T allele was found to confer a 1.6-fold risk for hypertension in women (95% confidence interval (CI), 1.132.06), whereas when combined with the effect of the ACE DD genotype, the risk was 2.34-fold (95% CI, 1.754.85) in two independent samples. The adjusted diastolic blood pressure response to Captopril was 3.3 mm Hg lower in ACE2 T allele carriers than in CC genotype carriers (P=0.019) in women. We conclude that the ACE2 T allele confers a high risk for hypertension and reduced antihypertensive response to ACE inhibitors.

  9. Indirect modulation of Shh signaling by Dlx5 affects the oral-nasal patterning of palate and rescues cleft palate in Msx1-null mice.

    PubMed

    Han, Jun; Mayo, Julie; Xu, Xun; Li, Jingyuan; Bringas, Pablo; Maas, Richard L; Rubenstein, John L R; Chai, Yang

    2009-12-01

    Cleft palate represents one of the most common congenital birth defects in human. During embryonic development, palatal shelves display oronasal (O-N) and anteroposterior polarity before the onset of fusion, but how the O-N pattern is established and how it relates to the expansion and fusion of the palatal shelves are unknown. Here we address these questions and show that O-N patterning is associated with the expansion and fusion of the palatal shelves and that Dlx5 is required for the O-N patterning of palatal mesenchyme. Loss of Dlx5 results in downregulation of Fgf7 and expanded Shh expression from the oral to the nasal side of the palatal shelf. This expanded Shh signaling is sufficient to restore palatal expansion and fusion in mice with compromised palatal mesenchymal cell proliferation, such as Msx1-null mutants. Exogenous Fgf7 inhibits Shh signaling and reverses the cranial neural crest (CNC) cell proliferation rescue in the Msx1/Dlx5 double knockout palatal mesenchyme. Thus, Dlx5-regulated Fgf7 signaling inhibits the expression of Shh, which in turn controls the fate of CNC cells through tissue-tissue interaction and plays a crucial role during palatogenesis. Our study shows that modulation of Shh signaling may be useful as a potential therapeutic approach for rescuing cleft palate.

  10. Noninvasive and real-time assessment of reconstructed functional human endometrium in NOD/SCID/gamma c(null) immunodeficient mice.

    PubMed

    Masuda, Hirotaka; Maruyama, Tetsuo; Hiratsu, Emi; Yamane, Junichi; Iwanami, Akio; Nagashima, Takashi; Ono, Masanori; Miyoshi, Hiroyuki; Okano, Hirotaka James; Ito, Mamoru; Tamaoki, Norikazu; Nomura, Tatsuji; Okano, Hideyuki; Matsuzaki, Yumi; Yoshimura, Yasunori

    2007-02-06

    Human uterine endometrium exhibits unique properties of cyclical regeneration and remodeling throughout reproductive life and also is subject to endometriosis through ectopic implantation of retrogradely shed endometrial fragments during menstruation. Here we show that functional endometrium can be regenerated from singly dispersed human endometrial cells transplanted beneath the kidney capsule of NOD/SCID/gamma(c)(null) immunodeficient mice. In addition to the endometrium-like structure, hormone-dependent changes, including proliferation, differentiation, and tissue breakdown and shedding (menstruation), can be reproduced in the reconstructed endometrium, the blood to which is supplied predominantly by human vessels invading into the mouse kidney parenchyma. Furthermore, the hormone-dependent behavior of the endometrium regenerated from lentivirally engineered endometrial cells expressing a variant luciferase can be assessed noninvasively and quantitatively by in vivo bioluminescence imaging. These results indicate that singly dispersed endometrial cells have potential applications for tissue reconstitution, angiogenesis, and human-mouse chimeric vessel formation, providing implications for mechanisms underlying the physiological endometrial regeneration during the menstrual cycle and the establishment of endometriotic lesions. This animal system can be applied as the unique model of endometriosis or for other various types of neoplastic diseases with the capacity of noninvasive and real-time evaluation of the effect of therapeutic agents and gene targeting when the relevant cells are transplanted beneath the kidney capsule.

  11. Quantitative Trait Loci Affecting Atherosclerosis at the Aortic Root Identified in an Intercross between DBA2J and 129S6 Apolipoprotein E-Null Mice

    PubMed Central

    Kayashima, Yukako; Tomita, Hirofumi; Zhilicheva, Svetlana; Kim, Shinja; Kim, Hyung-Suk; Bennett, Brian J.; Maeda, Nobuyo

    2014-01-01

    Apolipoprotein E-null mice on a DBA/2J genetic background (DBA-apoE) are highly susceptible to atherosclerosis in the aortic root area compared with those on a 129S6 background (129-apoE). To explore atherosclerosis-responsible genetic regions, we performed a quantitative trait locus (QTL) analysis using 172 male and 137 female F2 derived from an intercross between DBA-apoE and 129-apoE mice. A genome-wide scan identified two significant QTL for the size of lesions at the root: one is Ath44 on Chromosome (Chr) 1 at 158 Mb, and the other Ath45 on Chr 2 at 162 Mb. Ath44 co-localizes with but appears to be independent of a previously reported QTL, Ath1, while Ath45 is a novel QTL. DBA alleles of both Ath44 and Ath45 confer atherosclerosis-susceptibility. In addition, a QTL on Chr 14 at 73 Mb was found significant only in males, and 129 allele conferring susceptibility. Further analysis detected female-specific interactions between a second QTL on Chr 1 at 73 Mb and a QTL on Chr 3 at 21 Mb, and between Chr 7 at 84 Mb and Chr 12 at 77 Mb. These loci for the root atherosclerosis were independent of QTLs for plasma total cholesterol and QTLs for triglycerides, but a QTL for HDL (Chr 1 at 126 Mb) overlapped with the Ath44. Notably, haplotype analysis among 129S6, DBA/2J and C57BL/6 genomes and their gene expression data narrowed the candidate regions for Ath44 and Ath45 to less than 5 Mb intervals where multiple genome wide associations with cardiovascular phenotypes have also been reported in humans. SNPs in or near Fmo3, Sele and Selp for Ath44, and Lbp and Pkig for Ath45 were suggested for further investigation as potential candidates underlying the atherosclerosis susceptibility. PMID:24586312

  12. Null and hypomorph Prickle1 alleles in mice phenocopy human Robinow syndrome and disrupt signaling downstream of Wnt5a

    PubMed Central

    Liu, Chunqiao; Lin, Chen; Gao, Chun; May-Simera, Helen; Swaroop, Anand; Li, Tiansen

    2014-01-01

    ABSTRACT Planar cell polarity (PCP) signaling plays a critical role in tissue morphogenesis. In mammals, disruption of three of the six “core PCP” components results in polarity-dependent defects with rotated cochlear hair cell stereocilia and open neural tube. We recently demonstrated a role of Prickle1, a core PCP molecule in Drosophila, in mammalian neuronal development. To examine Prickle1 function along a broader developmental window, we generated three mutant alleles in mice. We show that the complete loss of Prickle1 leads to systemic tissue outgrowth defects, aberrant cell organization and disruption of polarity machinery. Curiously, Prickle1 mutants recapitulate the characteristic features of human Robinow syndrome and phenocopy mouse mutants with Wnt5a or Ror2 gene defects, prompting us to explore an association of Prickle1 with the Wnt pathway. We show that Prickle1 is a proteasomal target of Wnt5a signaling and that Dvl2, a target of Wnt5a signaling, is misregulated in Prickle1 mutants. Our studies implicate Prickle1 as a key component of the Wnt-signaling pathway and suggest that Prickle1 mediates some of the WNT5A-associated genetic defects in Robinow syndrome. PMID:25190059

  13. MMP20, KLK4, and MMP20/KLK4 double null mice define roles for matrix proteases during dental enamel formation.

    PubMed

    Hu, Yuanyuan; Smith, Charles E; Richardson, Amelia S; Bartlett, John D; Hu, Jan C C; Simmer, James P

    2016-03-01

    Matrix metalloproteinase 20 (MMP20) and kallikrein-related peptidase 4 (KLK4) are secreted proteinases that are essential for proper dental enamel formation. We characterized and compared enamel formed in wild-type, Mmp20 (-/-), Klk4 (-/-), Mmp20 (+/-) Klk4 (+/-), and Mmp20 (-/-) Klk4 (-/-) mice using dissecting and light microscopy, backscattered scanning electron microscopy (bSEM), SEM, microcomputed tomography (μCT), and energy-dispersive X-ray analysis (EDX). Following eruption, fractures were observed on Mmp20 (-/-), Klk4 (-/-), Mmp20 (+/-) Klk4 (+/-), and Mmp20 (-/-) Klk4 (-/-) molars. Failure of the enamel in the Mmp20 (+/-) Klk4 (+/-) molars was unexpected and suggested that digenic effects could contribute to the etiology of amelogenesis imperfecta in humans. Micro-CT analyses of hemimandibles demonstrated significantly reduced high-density enamel volume in the Mmp20 (-/-) and Klk4 (-/-) mice relative to the wild-type, which was further reduced in Mmp20 (-/-) Klk4 (-/-) mice. bSEM images of 7-week Mmp20 (-/-) and Mmp20 (-/-) Klk4 (-/-) mandibular incisors showed rough, pitted enamel surfaces with numerous indentations and protruding nodules. The Mmp20 (+/-) and Mmp20 (+/-) Klk4 (+/-) incisors showed prominent, evenly spaced, horizontal ridges that were more distinct in Mmp20 (+/-) Klk4 (+/-) incisors relative to Mmp20 (+/-) incisors due to the darkening of the valleys between the ridges. In cross sections, the Mmp20 (-/-) and Mmp20 (-/-) Klk4 (-/-) exhibited three distinct layers. The outer layer exhibited a disturbed elemental composition and an irregular enamel surface covered with nodules. The Mmp20 null enamel was apparently unable to withstand the sheer forces associated with eruption and separated from dentin during development. Cells invaded the cracks and interposed between the dentin and enamel layers. MMP20 and KLK4 serve overlapping and complementary functions to harden enamel by removing protein, but MMP20 potentially serves multiple

  14. Hyper-Variability in Circulating Insulin, High Fat Feeding Outcomes, and Effects of Reducing Ins2 Dosage in Male Ins1-Null Mice in a Specific Pathogen-Free Facility

    PubMed Central

    Templeman, Nicole M.; Mehran, Arya E.; Johnson, James D.

    2016-01-01

    Insulin is an essential hormone with key roles in energy homeostasis and body composition. Mice and rats, unlike other mammals, have two insulin genes: the rodent-specific Ins1 gene and the ancestral Ins2 gene. The relationships between insulin gene dosage and obesity has previously been explored in male and female Ins2-/- mice with full or reduced Ins1 dosage, as well as in female Ins1-/- mice with full or partial Ins2 dosage. We report herein unexpected hyper-variability in Ins1-null male mice, with respect to their circulating insulin levels and to the physiological effects of modulating Ins2 gene dosage. Two large cohorts of Ins1-/-:Ins2+/- mice and their Ins1-/-:Ins2+/+ littermates were fed chow diet or high fat diet (HFD) from weaning, and housed in specific pathogen-free conditions. Cohort A and cohort B were studied one year apart. Contrary to female mice from the same litters, inactivating one Ins2 allele on the complete Ins1-null background did not consistently cause a reduction of circulating insulin in male mice, on either diet. In cohort A, all HFD-fed males showed an equivalent degree of insulin hypersecretion and weight gain, regardless of Ins2 dosage. In cohort B the effects of HFD appeared generally diminished, and cohort B Ins1-/-:Ins2+/- males showed decreased insulin levels and body mass compared to Ins1-/-:Ins2+/+ littermates, on both diets. Although experimental conditions were consistent between cohorts, we found that HFD-fed Ins1-/-:Ins2+/- mice with lower insulin levels had increased corticosterone. Collectively, these observations highlight the phenotypic characteristics that change in association with differences in circulating insulin and Ins2 gene dosage, particularly in male mice. PMID:27055260

  15. Angiotensin-converting enzyme 2 amplification limited to the circulation does not protect mice from development of diabetic nephropathy.

    PubMed

    Wysocki, Jan; Ye, Minghao; Khattab, Ahmed M; Fogo, Agnes; Martin, Aline; David, Nicolae Valentin; Kanwar, Yashpal; Osborn, Mark; Batlle, Daniel

    2016-12-04

    Blockers of the renin-angiotensin system are effective in the treatment of experimental and clinical diabetic nephropathy. An approach different from blocking the formation or action of angiotensin II (1-8) that could also be effective involves fostering its degradation. Angiotensin-converting enzyme 2 (ACE2) is a monocarboxypeptidase that cleaves angiotensin II (1-8) to form angiotensin (1-7). Therefore, we examined the renal effects of murine recombinant ACE2 in mice with streptozotocin-induced diabetic nephropathy as well as that of amplification of circulating ACE2 using minicircle DNA delivery prior to induction of experimental diabetes. This delivery resulted in a long-term sustained and profound increase in serum ACE2 activity and enhanced ability to metabolize an acute angiotensin II (1-8) load. In mice with streptozotocin-induced diabetes pretreated with minicircle ACE2, ACE2 protein in plasma increased markedly and this was associated with a more than 100-fold increase in serum ACE2 activity. However, minicircle ACE2 did not result in changes in urinary ACE2 activity as compared to untreated diabetic mice. In both diabetic groups, glomerular filtration rate increased significantly and to the same extent as compared to non-diabetic controls. Albuminuria, glomerular mesangial expansion, glomerular cellularity, and glomerular size were all increased to a similar extent in minicircle ACE2-treated and untreated diabetic mice, as compared to non-diabetic controls. Recombinant mouse ACE2 given for 4 weeks by intraperitoneal daily injections in mice with streptozotocin-induced diabetic nephropathy also failed to improve albuminuria or kidney pathology. Thus, a profound augmentation of ACE2 confined to the circulation failed to ameliorate the glomerular lesions and hyperfiltration characteristic of early diabetic nephropathy. These findings emphasize the importance of targeting the kidney rather than the circulatory renin angiotensin system to combat diabetic

  16. Shipboard Sunphotometer Measurements of Aerosol Optical Depth During ACE-2 and Comparison with Selected Ship, Aircraft and Satellite Measurements

    NASA Technical Reports Server (NTRS)

    Livingston, J. M.; Kapustin, V. N.; Schmid, B.; Russell, P. B.; Quinn, P. K.; Bates, T. S.; Durkee, P. A.; Nielsen, K.; Freudenthaler, V.; Wiegner, M.; Covert, D. S.

    2000-01-01

    We present analyses of aerosol optical depth (AOD) measurements taken with a shipboard six-channel tracking sunphotometer during ACE-2. For 10 July 1997, results are also shown for measurements acquired 70 km from the ship with a fourteen-channel airborne tracking sunphotometer.

  17. North Atlantic Aerosol Properties for Radiative Impact Assessments. Derived from Column Closure Analyses in TARFOX and ACE-2

    NASA Technical Reports Server (NTRS)

    Russell, Philip A.; Bergstrom, Robert A.; Schmid, Beat; Livingston, John M.

    2000-01-01

    Aerosol effects on atmospheric radiative fluxes provide a forcing function that can change the climate in potentially significant ways. This aerosol radiative forcing is a major source of uncertainty in understanding the climate change of the past century and predicting future climate. To help reduce this uncertainty, the 1996 Tropospheric Aerosol Radiative Forcing Observational Experiment (TARFOX) and the 1997 Aerosol Characterization Experiment (ACE-2) measured the properties and radiative effects of aerosols over the Atlantic Ocean. Both experiments used remote and in situ measurements from aircraft and the surface, coordinated with overpasses by a variety of satellite radiometers. TARFOX focused on the urban-industrial haze plume flowing from the United States over the western Atlantic, whereas ACE-2 studied aerosols over the eastern Atlantic from both Europe and Africa. These aerosols often have a marked impact on satellite-measured radiances. However, accurate derivation of flux changes, or radiative forcing, from the satellite measured radiances or retrieved aerosol optical depths (AODs) remains a difficult challenge. Here we summarize key initial results from TARFOX and ACE-2, with a focus on closure analyses that yield aerosol microphysical models for use in improved assessments of flux changes. We show how one such model gives computed radiative flux sensitivities (dF/dAOD) that agree with values measured in TARFOX and preliminary values computed for the polluted marine boundary layer in ACE-2. A companion paper uses the model to compute aerosol-induced flux changes over the North Atlantic from AVHRR-derived AOD fields.

  18. BDNF-Mediated Cerebellar Granule Cell Development is Impaired in Mice Null for CaMKK2 or CaMKIV

    PubMed Central

    Kokubo, Manabu; Nishio, Masahiro; Ribar, Thomas J.; Anderson, Kristin A.; West, Anne E.; Means, Anthony R.

    2009-01-01

    The Ca2+/calmodulin-activated kinases CamKK2 and CaMKIV are highly expressed in the brain where they play important roles in activating intracellular responses to elevated Ca2+. To address the biological functions of Ca2+ signaling via these kinases during brain development we have examined cerebellar development in mice null for CaMKK2 or CaMKIV. Here we demonstrate that CaMKK2/CaMKIV-dependent phosphorylation of CREB correlates with Bdnf transcription, which is required for normal development of cerebellar granule cell neurons. We show in vivo and in vitro that the absence of either CaMKK2 or CaMKIV disrupts the ability of developing cerebellar granule cells in the external granule cell layer to cease proliferation and begin migration to the internal granule cell layer. Further, loss of CaMKK2 or CaMKIV results in decreased pCREB, Bdnf exon I and IV-containing mRNAs and BDNF protein in cerebellar granule cell neurons. Re-expression of CaMKK2 or CaMKIV in granule cells that lack CaMKK2 or CaMKIV, respectively, restores pCREB and BDNF to wild type levels and addition of BDNF rescues granule cell migration in vitro. These results reveal a previously undefined role for a CaMKK2/CaMKIV cascade involved in cerebellar granule cell development and show specifically that Ca2+-dependent regulation of BDNF through CaMKK2/CaMKIV is required for this process. PMID:19605628

  19. ILDR1 null mice, a model of human deafness DFNB42, show structural aberrations of tricellular tight junctions and degeneration of auditory hair cells.

    PubMed

    Morozko, Eva L; Nishio, Ayako; Ingham, Neil J; Chandra, Rashmi; Fitzgerald, Tracy; Martelletti, Elisa; Borck, Guntram; Wilson, Elizabeth; Riordan, Gavin P; Wangemann, Philine; Forge, Andrew; Steel, Karen P; Liddle, Rodger A; Friedman, Thomas B; Belyantseva, Inna A

    2015-02-01

    In the mammalian inner ear, bicellular and tricellular tight junctions (tTJs) seal the paracellular space between epithelial cells. Tricellulin and immunoglobulin-like (Ig-like) domain containing receptor 1 (ILDR1, also referred to as angulin-2) localize to tTJs of the sensory and non-sensory epithelia in the organ of Corti and vestibular end organs. Recessive mutations of TRIC (DFNB49) encoding tricellulin and ILDR1 (DFNB42) cause human nonsyndromic deafness. However, the pathophysiology of DFNB42 deafness remains unknown. ILDR1 was recently reported to be a lipoprotein receptor mediating the secretion of the fat-stimulated cholecystokinin (CCK) hormone in the small intestine, while ILDR1 in EpH4 mouse mammary epithelial cells in vitro was shown to recruit tricellulin to tTJs. Here we show that two different mouse Ildr1 mutant alleles have early-onset severe deafness associated with a rapid degeneration of cochlear hair cells (HCs) but have a normal endocochlear potential. ILDR1 is not required for recruitment of tricellulin to tTJs in the cochlea in vivo; however, tricellulin becomes mislocalized in the inner ear sensory epithelia of ILDR1 null mice after the first postnatal week. As revealed by freeze-fracture electron microscopy, ILDR1 contributes to the ultrastructure of inner ear tTJs. Taken together, our data provide insight into the pathophysiology of human DFNB42 deafness and demonstrate that ILDR1 is crucial for normal hearing by maintaining the structural and functional integrity of tTJs, which are critical for the survival of auditory neurosensory HCs.

  20. Peptide Mimetic of the S100A4 Protein Modulates Peripheral Nerve Regeneration and Attenuates the Progression of Neuropathy in Myelin Protein P0 Null Mice

    PubMed Central

    Moldovan, Mihai; Pinchenko, Volodymyr; Dmytriyeva, Oksana; Pankratova, Stanislava; Fugleholm, Kåre; Klingelhofer, Jorg; Bock, Elisabeth; Berezin, Vladimir; Krarup, Christian; Kiryushko, Darya

    2013-01-01

    We recently found that S100A4, a member of the multifunctional S100 protein family, protects neurons in the injured brain and identified two sequence motifs in S100A4 mediating its neurotrophic effect. Synthetic peptides encompassing these motifs stimulated neuritogenesis and survival in vitro and mimicked the S100A4-induced neuroprotection in brain trauma. Here, we investigated a possible function of S100A4 and its mimetics in the pathologies of the peripheral nervous system (PNS). We found that S100A4 was expressed in the injured PNS and that its peptide mimetic (H3) affected the regeneration and survival of myelinated axons. H3 accelerated electrophysiological, behavioral and morphological recovery after sciatic nerve crush while transiently delaying regeneration after sciatic nerve transection and repair. On the basis of the finding that both S100A4 and H3 increased neurite branching in vitro, these effects were attributed to the modulatory effect of H3 on initial axonal sprouting. In contrast to the modest effect of H3 on the time course of regeneration, H3 had a long-term neuroprotective effect in the myelin protein P0 null mice, a model of dysmyelinating neuropathy (Charcot-Marie-Tooth type 1 disease), where the peptide attenuated the deterioration of nerve conduction, demyelination and axonal loss. From these results, S100A4 mimetics emerge as a possible means to enhance axonal sprouting and survival, especially in the context of demyelinating neuropathies with secondary axonal loss, such as Charcot-Marie-Tooth type 1 disease. Moreover, our data suggest that S100A4 is a neuroprotectant in PNS and that other S100 proteins, sharing high homology in the H3 motif, may have important functions in PNS pathologies. PMID:23508572

  1. Structural determinants for binding to angiotensin converting enzyme 2 (ACE2) and angiotensin receptors 1 and 2

    PubMed Central

    Clayton, Daniel; Hanchapola, Iresha; Thomas, Walter G.; Widdop, Robert E.; Smith, Alexander I.; Perlmutter, Patrick; Aguilar, Marie-Isabel

    2015-01-01

    Angiotensin converting enzyme 2 (ACE2) is a zinc carboxypeptidase involved in the renin–angiotensin system (RAS) and inactivates the potent vasopressive peptide angiotensin II (Ang II) by removing the C-terminal phenylalanine residue to yield Ang1–7. This conversion inactivates the vasoconstrictive action of Ang II and yields a peptide that acts as a vasodilatory molecule at the Mas receptor and potentially other receptors. Given the growing complexity of RAS and level of cross-talk between ligands and their corresponding enzymes and receptors, the design of molecules with selectivity for the major RAS binding partners to control cardiovascular tone is an on-going challenge. In previous studies we used single β-amino acid substitutions to modulate the structure of Ang II and its selectivity for ACE2, AT1R, and angiotensin type 2 (AT2R) receptor. We showed that modification at the C-terminus of Ang II generally resulted in more pronounced changes to secondary structure and ligand binding, and here, we further explore this region for the potential to modulate ligand specificity. In this study, (1) a library of 47 peptides derived from the C-terminal tetrapeptide sequence (-IHPF) of Ang II was synthesized and assessed for ACE2 binding, (2) the terminal group requirements for high affinity ACE2 binding were explored by and N- and C-terminal modification, (3) high affinity ACE2 binding chimeric AngII analogs were then synthesized and assessed, (4) the structure of the full-length Ang II analogs were assessed by circular dichroism, and (5) the Ang II analogs were assessed for AT1R/AT2R selectivity by cell-based assays. Studies on the C-terminus of Ang II demonstrated varied specificity at different residue positions for ACE2 binding and four Ang II chimeric peptides were identified as selective ligands for the AT2 receptor. Overall, these results provide insight into the residue and structural requirements for ACE2 binding and angiotensin receptor selectivity. PMID

  2. ACE2/Ang-(1-7)/Mas axis stimulates vascular repair-relevant functions of CD34+ cells.

    PubMed

    Singh, Neha; Joshi, Shrinidh; Guo, Lirong; Baker, Matthew B; Li, Yan; Castellano, Ronald K; Raizada, Mohan K; Jarajapu, Yagna P R

    2015-11-15

    CD34(+) stem/progenitor cells have been identified as a promising cell population for the autologous cell-based therapies in patients with cardiovascular disease. The counter-regulatory axes of renin angiotensin system, angiotensin converting enzyme (ACE)/Ang II/angiotensin type 1 (AT1) receptor and ACE2/Ang-(1-7)/Mas receptor, play an important role in the cardiovascular repair. This study evaluated the expression and vascular repair-relevant functions of these two pathways in human CD34(+) cells. CD34(+) cells were isolated from peripheral blood mononuclear cells (MNCs), obtained from healthy volunteers. Expression of ACE, ACE2, AT1, and angiotensin type 2 and Mas receptors were determined. Effects of Ang II, Ang-(1-7), Norleu(3)-Ang-(1-7), and ACE2 activators, xanthenone (XNT) and diminazene aceturate (DIZE) on proliferation, migration, and adhesion of CD34(+) cells were evaluated. ACE2 and Mas were relatively highly expressed in CD34(+) cells compared with MNCs. Ang-(1-7) or its analog, Norleu(3)-Ang-(1-7), stimulated proliferation of CD34(+) cells that was associated with decrease in phosphatase and tensin homologue deleted on chromosome 10 levels and was inhibited by triciribin, an AKT inhibitor. Migration of CD34(+) cells was enhanced by Ang-(1-7) or Norleu(3)-Ang-(1-7) that was decreased by a Rho-kinase inhibitor, Y-27632. In the presence of Ang II, XNT or DIZE enhanced proliferation and migration that were blocked by DX-600, an ACE2 inhibitor. Treatment of MNCs with Ang II, before the isolation of CD34(+) cells, attenuated the proliferation and migration to stromal derived factor-1α. This attenuation was reversed by apocynin, an NADPH oxidase inhibitor. Adhesion of MNCs or CD34(+) cells to fibronectin was enhanced by Ang II and was unaffected by Ang-(1-7). This study suggests that ACE2/Ang-(1-7)/Mas pathway stimulates functions of CD34(+) cells that are cardiovascular protective, whereas Ang II attenuates these functions by acting on MNCs. These findings

  3. Isolation and characterization of a bat SARS-like coronavirus that uses the ACE2 receptor

    PubMed Central

    Ge, Xing-Yi; Li, Jia-Lu; Yang, Xing-Lou; Chmura, Aleksei A.; Zhu, Guangjian; Epstein, Jonathan H.; Mazet, Jonna K.; Hu, Ben; Zhang, Wei; Peng, Cheng; Zhang, Yu-Ji; Luo, Chu-Ming; Tan, Bing; Wang, Ning; Zhu, Yan; Crameri, Gary; Zhang, Shu-Yi; Wang, Lin-Fa; Daszak, Peter; Shi, Zheng-Li

    2016-01-01

    The 2002–3 pandemic caused by severe acute respiratory syndrome coronavirus (SARS-CoV) was one of the most significant public health events in recent history1. An ongoing outbreak of Middle East respiratory syndrome coronavirus (MERS-CoV)2 suggests that this group of viruses remains a major threat and that their distribution is wider than previously recognized. Although bats have been suggested as the natural reservoirs of both viruses3–5, attempts to isolate the progenitor virus of SARS-CoV from bats have been unsuccessful. Diverse SARS-like coronaviruses (SL-CoVs) have now been reported from bats in China, Europe and Africa5–8, but none are considered a direct progenitor of SARS-CoV because of their phylogenetic disparity from this virus and the inability of their spike proteins (S) to use the SARS-CoV cellular receptor molecule, the human angiotensin converting enzyme II (ACE2)9,10. Here, we report whole genome sequences of two novel bat CoVs from Chinese horseshoe bats (Family: Rhinolophidae) in Yunnan, China; RsSHC014 and Rs3367. These viruses are far more closely related to SARS-CoV than any previously identified bat CoVs, particularly in the receptor binding domain (RDB) of the S protein. Most importantly, we report the first recorded isolation of a live SL-CoV (bat SL-CoV-WIV1) from bat fecal samples in Vero E6 cells, which has typical coronavirus morphology, 99.9% sequence identity to Rs3367 and uses the ACE2s from human, civet and Chinese horseshoe bat for cell entry. Preliminary in vitro testing indicates that WIV1 also has a broad species tropism. Our results provide the strongest evidence to date that Chinese horseshoe bats are natural reservoirs of SARS-CoV, and that intermediate hosts may not be necessary for direct human infection by some bat SL-CoVs. They also highlight the importance of pathogen discovery programs targeting high-risk wildlife groups in emerging disease hotspots as a strategy for pandemic preparedness. PMID:24172901

  4. Isolation and characterization of a bat SARS-like coronavirus that uses the ACE2 receptor.

    PubMed

    Ge, Xing-Yi; Li, Jia-Lu; Yang, Xing-Lou; Chmura, Aleksei A; Zhu, Guangjian; Epstein, Jonathan H; Mazet, Jonna K; Hu, Ben; Zhang, Wei; Peng, Cheng; Zhang, Yu-Ji; Luo, Chu-Ming; Tan, Bing; Wang, Ning; Zhu, Yan; Crameri, Gary; Zhang, Shu-Yi; Wang, Lin-Fa; Daszak, Peter; Shi, Zheng-Li

    2013-11-28

    The 2002-3 pandemic caused by severe acute respiratory syndrome coronavirus (SARS-CoV) was one of the most significant public health events in recent history. An ongoing outbreak of Middle East respiratory syndrome coronavirus suggests that this group of viruses remains a key threat and that their distribution is wider than previously recognized. Although bats have been suggested to be the natural reservoirs of both viruses, attempts to isolate the progenitor virus of SARS-CoV from bats have been unsuccessful. Diverse SARS-like coronaviruses (SL-CoVs) have now been reported from bats in China, Europe and Africa, but none is considered a direct progenitor of SARS-CoV because of their phylogenetic disparity from this virus and the inability of their spike proteins to use the SARS-CoV cellular receptor molecule, the human angiotensin converting enzyme II (ACE2). Here we report whole-genome sequences of two novel bat coronaviruses from Chinese horseshoe bats (family: Rhinolophidae) in Yunnan, China: RsSHC014 and Rs3367. These viruses are far more closely related to SARS-CoV than any previously identified bat coronaviruses, particularly in the receptor binding domain of the spike protein. Most importantly, we report the first recorded isolation of a live SL-CoV (bat SL-CoV-WIV1) from bat faecal samples in Vero E6 cells, which has typical coronavirus morphology, 99.9% sequence identity to Rs3367 and uses ACE2 from humans, civets and Chinese horseshoe bats for cell entry. Preliminary in vitro testing indicates that WIV1 also has a broad species tropism. Our results provide the strongest evidence to date that Chinese horseshoe bats are natural reservoirs of SARS-CoV, and that intermediate hosts may not be necessary for direct human infection by some bat SL-CoVs. They also highlight the importance of pathogen-discovery programs targeting high-risk wildlife groups in emerging disease hotspots as a strategy for pandemic preparedness.

  5. The heparan sulphate deficient Hspg2 exon 3 null mouse displays reduced deposition of TGF-β1 in skin compared to C57BL/6 wild type mice.

    PubMed

    Shu, Cindy; Smith, Susan M; Melrose, James

    2016-06-01

    This was an observational study where we examined the role of perlecan HS on the deposition of TGF-β1 in C57BL/6 and Hspg2(∆3-/∆3-) perlecan exon 3 null mouse skin. Despite its obvious importance in skin repair and tissue homeostasis no definitive studies have immunolocalised TGF-β1 in skin in WT or Hspg2(∆3-/∆3-) perlecan exon 3 null mice. Vertical parasagittal murine dorsal skin from 3, 6 and 12 week old C57BL/6 and Hspg2(∆3-/∆3-) mice were fixed in neutral buffered formalin, paraffin embedded and 4 μm sections stained with Mayers haematoxylin and eosin (H & E). TGF-β1 was immunolocalised using a rabbit polyclonal antibody, heat retrieval and the Envision NovaRED detection system. Immunolocalisation of TGF-β1 differed markedly in C57BL/6 and Hspg2(∆3-/∆3-) mouse skin, ablation of exon 3 of Hspg2 resulted in a very severe reduction in the deposition of TGF-β1 in skin 3-12 weeks postnatally. The reduced deposition of TGF-β1 observed in the present study would be expected to impact detrimentally on the remodelling and healing capacity of skin in mutant mice compounding on the poor wound-healing properties already reported for perlecan exon 3 null mice due to an inability to signal with FGF-2 and promote angiogenic repair processes. TGF-β1 also has cell mediated effects in tissue homeostasis and matrix stabilisation a reduction in TGF-β1 deposition would therefore be expected to detrimentally impact on skin homeostasis in the perlecan mutant mice.

  6. Relationship between hemodynamics and atherosclerosis in aortic arches of apolipoprotein E-null mice on 129S6/SvEvTac and C57BL/6J genetic backgrounds

    PubMed Central

    Tomita, Hirofumi; Hagaman, John; Friedman, Morton H.; Maeda, Nobuyo

    2011-01-01

    Objective We investigated the relationships between hemodynamics and differential plaque development at the aortic arch of apolipoprotein E (apoE)-null mice on 129S6/SvEvTac (129) and C57BL/6J (B6) genetic backgrounds. Methods Mean flow velocities at the ascending and descending aorta (mVAA and mVDA) were measured by Doppler ultrasound in wild type and apoE-null male mice at 3 and 9 months of age. Following dissection of the aortic arches, anatomical parameters and plaque areas were evaluated. Results Arch plaques were five times bigger in 129-apoE than in B6-apoE mice at 3 months, and twice as large at 9 months. The geometric differences, namely larger vessel diameter in the B6 strain and broader inner curvature of the aortic arch in the 129 strain, were exaggerated in 9-month-old apoE-null mice. Cardiac output and heart rate under anesthesia were significantly higher in the B6 strain than in the 129 strain. The values of mVAA were similar in the two strains, while mVDA was lower in the 129 strain. However, there was a 129-apoE-specific reduction of flow velocities with age, and both mVAA and mVDA were significantly lower in 129-apoE than in B6-apoE mice at 9 months. The mean relative wall shear stress (rWSS) over the aortic arch in 129-apoE and B6-apoE mice were not different, but animals with lower mean rWSS had larger arch plaques within each strain. Conclusions The plaque formation in the arch of apoE-null mice is accompanied by strain-dependent changes in both arch geometry and hemodynamics. While arch plaque sizes negatively correlate with mean rWSS, additional factors are necessary to account for the strain differences in arch plaque development. PMID:22078246

  7. Genome duplication and mutations in ACE2 cause multicellular, fast-sedimenting phenotypes in evolved Saccharomyces cerevisiae

    PubMed Central

    Oud, Bart; Guadalupe-Medina, Victor; Nijkamp, Jurgen F.; de Ridder, Dick; Pronk, Jack T.; van Maris, Antonius J. A.; Daran, Jean-Marc

    2013-01-01

    Laboratory evolution of the yeast Saccharomyces cerevisiae in bioreactor batch cultures yielded variants that grow as multicellular, fast-sedimenting clusters. Knowledge of the molecular basis of this phenomenon may contribute to the understanding of natural evolution of multicellularity and to manipulating cell sedimentation in laboratory and industrial applications of S. cerevisiae. Multicellular, fast-sedimenting lineages obtained from a haploid S. cerevisiae strain in two independent evolution experiments were analyzed by whole genome resequencing. The two evolved cell lines showed different frameshift mutations in a stretch of eight adenosines in ACE2, which encodes a transcriptional regulator involved in cell cycle control and mother-daughter cell separation. Introduction of the two ace2 mutant alleles into the haploid parental strain led to slow-sedimenting cell clusters that consisted of just a few cells, thus representing only a partial reconstruction of the evolved phenotype. In addition to single-nucleotide mutations, a whole-genome duplication event had occurred in both evolved multicellular strains. Construction of a diploid reference strain with two mutant ace2 alleles led to complete reconstruction of the multicellular-fast sedimenting phenotype. This study shows that whole-genome duplication and a frameshift mutation in ACE2 are sufficient to generate a fast-sedimenting, multicellular phenotype in S. cerevisiae. The nature of the ace2 mutations and their occurrence in two independent evolution experiments encompassing fewer than 500 generations of selective growth suggest that switching between unicellular and multicellular phenotypes may be relevant for competitiveness of S. cerevisiae in natural environments. PMID:24145419

  8. Perinatally administered losartan augments renal ACE2 expression but not cardiac or renal Mas receptor in spontaneously hypertensive rats.

    PubMed

    Klimas, Jan; Olvedy, Michael; Ochodnicka-Mackovicova, Katarina; Kruzliak, Peter; Cacanyiova, Sona; Kristek, Frantisek; Krenek, Peter; Ochodnicky, Peter

    2015-08-01

    Since the identification of the alternative angiotensin converting enzyme (ACE)2/Ang-(1-7)/Mas receptor axis, renin-angiotensin system (RAS) is a new complex target for a pharmacological intervention. We investigated the expression of RAS components in the heart and kidney during the development of hypertension and its perinatal treatment with losartan in young spontaneously hypertensive rats (SHR). Expressions of RAS genes were studied by the RT-PCR in the left ventricle and kidney of rats: normotensive Wistar, untreated SHR, SHR treated with losartan since perinatal period until week 9 of age (20 mg/kg/day) and SHR treated with losartan only until week 4 of age and discontinued until week 9. In the hypertrophied left ventricle of SHR, cardiac expressions of Ace and Mas were decreased while those of AT1 receptor (Agtr1a) and Ace2 were unchanged. Continuous losartan administration reduced LV weight (0.43 ± 0.02; P < 0.05 versus SHR) but did not influence altered cardiac RAS expression. Increased blood pressure in SHR (149 ± 2 in SHR versus 109 ± 2 mmHg in Wistar; P < 0.05) was associated with a lower renal expressions of renin, Agtr1a and Mas and with an increase in ACE2. Continuous losartan administration lowered blood pressure to control levels (105 ± 3 mmHg; P < 0.05 versus SHR), however, only renal renin and ACE2 were significantly up-regulated (for both P < 0.05 versus SHR). Conclusively, prevention of hypertension and LV hypertrophy development by losartan was unrelated to cardiac or renal expression of Mas. Increased renal Ace2, and its further increase by losartan suggests the influence of locally generated Ang-(1-7) in organ response to the developing hypertension in SHRs.

  9. MPD in Telomerase Null Mice

    DTIC Science & Technology

    2007-09-01

    mechanisms and genetic pathways involved in the pathogenesis of myeloproliferative disease (MPDs) are not well understood. Telomere maintenance and...24). A large body of evidence demonstrates the importance of tyrosine kinase mutations to the development of myeloproliferative disorders

  10. Collectrin, a homologue of ACE2, its transcriptional control and functional perspectives

    SciTech Connect

    Zhang Yanling Wada, Jun

    2007-11-09

    Collectrin is a type I membrane protein and shares significant homology with C-terminal domain of angiotensin-converting enzyme-2 (ACE2). However, collectrin lacks catalytic domain and it suggests the presence of uncharacterized physiological functions of collectrin. Collectrin is transcriptionally regulated by hepatocyte nuclear factor-{alpha} and -{beta} and is highly expressed on renal proximal tubules and collecting ducts as well as pancreatic {beta}-cells. Recent in vitro and in vivo studies demonstrated interesting physiological roles of collectrin related to insulin secretion, formation of primary cilia, renal cyst formation and amino acid transport. The common underlying molecular mechanism may be suggested by the evidence that collectrin binds to SNARE complex by interacting with snapin. Collectrin is involved in the process of vesicle transport and membrane fusion and thus it delivers insulin for exocytosis or various membrane proteins to apical plasmalemma and primary cilia. Collectrin may be the new therapeutic target for various pathological processes such as diabetes, polycystic kidney disease, hypertension and aminoaciduria.

  11. Effects of felodipine combined with puerarin on ACE2-Ang (1-7)-Mas axis in renovascular hypertensive rat.

    PubMed

    Bai, Song; Huang, Zheng-Gui; Chen, Li; Wang, Jiang-Tao; Ding, Bo-Ping

    2013-06-10

    This study aimed to investigate the effect of combination of felodipine+puerarin on ACE2-Ang (1-7)-Mas axis, and to explore the protective effect of the combination against kidney in renovascular hypertensive rats. Goldblatt rats were randomly divided into 5 groups as follows: 4 groups which were treated with felodipine (Felo), puerarin (Pue), Felo+Pue, and Felo+captopril (Cap), respectively, and a control group of animals that were administrated with distilled water. Contents of Ang II and Ang (1-7) in renal tissues were determined by ELISA kit. The mRNA expression of ACE2/Mas and ACE/AT1 in kidneys was analyzed by RT-PCR. After 8weeks of treatment, compared with Goldblatt group, Felo+Pue reduced SBP, DBP and HR (p<0.01 or p<0.05), ameliorated renal interstitial fibrosis, decreased the level of Ang II and increased that of Ang (1-7), upregulated mRNA expression of ACE2 and Mas, decreased that of ACE and AT1, and downregulated protein expression of TGF-β1 in kidneys (p<0.01). Compared with Felo group, Felo+Pue decreased DBP and HR more markedly, attenuated fibrosis, decreased Ang II levels and increased those of Ang (1-7), upregulated mRNA expression of ACE2 in bilateral kidneys and that of Mas in ischemic kidney, downregulated that of ACE in bilateral kidneys and that of AT1 in ischemic kidney, and decreased expression of TGF-β1 protein significantly. In a word, a combination of Felo+Pue has a more efficient therapeutic effect on DBP and HR, and contributes to a better protection against renal interstitial fibrosis.

  12. Captopril improves postresuscitation hemodynamics protective against pulmonary embolism by activating the ACE2/Ang-(1-7)/Mas axis.

    PubMed

    Xiao, Hong-Li; Li, Chun-Sheng; Zhao, Lian-Xing; Yang, Jun; Tong, Nan; An, Le; Liu, Qi-Tong

    2016-11-01

    Acute pulmonary embolism (APE) has a very high mortality rate, especially at cardiac arrest and even after the return of spontaneous circulation (ROSC). This study investigated the protective effect of the angiotensin-converting enzyme (ACE) inhibitor captopril on postresuscitation hemodynamics, in a porcine model of cardiac arrest established by APE. Twenty-nine Beijing Landrace pigs were infused with an autologous thrombus leading to cardiac arrest and subjected to standard cardiopulmonary resuscitation and thrombolysis. Ten resuscitated pigs were randomly and equally apportioned to receive either captopril (22.22 mg/kg) infusion or the same volume saline, 30 min after ROSC. Hemodynamic changes and ACE-Ang II-angiotensin II type 1 receptor (AT1R) and ACE2/Ang-(1-7)/Mas receptor axis levels were determined. APE was associated with a decline in mean arterial pressure and a dramatic increase in pulmonary artery pressure and mean right ventricular pressure. After ROSC, captopril infusion was associated with significantly lower mean right ventricular pressure and systemic and pulmonary vascular resistance, faster heart rate, and higher Ang-(1-7) levels, ACE2/ACE, and Ang-(1-7)/Ang II, compared with the saline infusion. The ACE2/Ang-(1-7)/Mas pathway correlated negatively with external vascular lung water and pulmonary vascular permeability and positively with the right cardiac index. In conclusion, in a pig model of APE leading to cardiac arrest, captopril infusion was associated with less mean right ventricular pressure overload after resuscitation, compared with saline infusion. The reduction in systemic and pulmonary vascular resistance associated with captopril may be by inhibiting the ACE-Ang II-AT1R axis and activating the ACE2/Ang-(1-7)/Mas axis.

  13. A novel role for PHT1 in the disposition of l-histidine in brain: In vitro slice and in vivo pharmacokinetic studies in wildtype and Pht1 null mice.

    PubMed

    Wang, Xiao-Xing; Hu, Yongjun; Keep, Richard F; Toyama-Sorimachi, Noriko; Smith, David E

    2017-01-15

    PHT1 (SLC15A4) is responsible for translocating l-histidine (l-His), di/tripeptides and peptide-like drugs across biological membranes. Previous studies have indicated that PHT1 is located in brain parenchyma, however, its role and significance in brain along with effect on the biodistribution of substrates is unknown. In this study, adult gender-matched Pht1-competent (wildtype) and Pht1-deficient (null) mice were used to investigate the effect of PHT1 on l-His brain disposition via in vitro slice and in vivo pharmacokinetic approaches. We also evaluated the serum clinical chemistry and expression levels of select transporters and enzymes in the two genotypes. No significant differences were observed between genotypes in serum chemistry, body weight, viability and fertility. PCR analyses indicated that Pept2 had a compensatory up-regulation in Pht1 null mice (about 2-fold) as compared to wildtype animals, which was consistent in different brain regions and confirmed by immunoblots. The uptake of l-His was reduced in brain slices by 50% during PHT1 ablation. The l-amino acid transporters accounted for 30% of the uptake, and passive (other) pathways for 20% of the uptake. During the in vivo pharmacokinetic studies, plasma concentration-time profiles of l-His were comparable between the two genotypes after intravenous administration. Still, biodistribution studies revealed that, when sampled 5min after dosing, l-His values were 28-48% lower in Pht1 null mice, as compared to wildtype animals, in brain parenchyma but not cerebrospinal fluid. These findings suggest that PHT1 may play an important role in histidine transport in brain, and resultant effects on histidine/histamine homeostasis and neuropeptide regulation.

  14. Hodgkin's lymphoma cells are efficiently engrafted and tumor marker CD30 is expressed with constitutive nuclear factor-kappaB activity in unconditioned NOD/SCID/gammac(null) mice.

    PubMed

    Dewan, Md Zahidunnabi; Watanabe, Mariko; Ahmed, Sunjida; Terashima, Kazuo; Horiuchi, Sankichi; Sata, Tetsutaro; Honda, Mitsuo; Ito, Mamoru; Watanabe, Toshiki; Horie, Ryouichi; Yamamoto, Naoki

    2005-08-01

    As there are very few reproducible animal models without conditioning available for the study of human B-cell-type Hodgkin's lymphoma (HL), we investigated the ability of HL cells to induce tumors using novel NOD/SCID/gammac(null) (NOG) mice. Four human Epstein-Barr virus-negative cell lines (KM-H2 and L428 originated from B cells, L540 and HDLM2 originated from T cells) were inoculated either subcutaneously in the postauricular region or intravenously in the tail of unmanipulated NOG mice. All cell lines successfully engrafted and produced tumors with infiltration of cells in various organs of all mice. Tumor cells had classical histomorphology as well as expression patterns of the tumor marker CD30, which is a cell surface antigen expressed on HL. Tumor progression in mice inoculated with B-cell-type, but not T-cell-type, HL cells correlated with an elevation in serum human interleukin-6 levels. Tumor cells from the mice also retained strong nuclear factor (NF)-kappaB DNA binding activity, and the induced NF-kappaB components were indistinguishable from those cultured in vitro. The reproducible growth behavior and preservation of characteristic features of both B-cell-type and T-cell-type HL in the mice suggest that this new xenotransplant model can provide a unique opportunity to understand and investigate the mechanism of pathogenesis and malignant cell growth, and to develop novel anticancer therapies.

  15. A Role for Endogenous Transforming Growth Factor β1 in Langerhans Cell Biology:  The Skin of   Transforming Growth Factor β1 Null Mice Is Devoid of  Epidermal Langerhans Cells

    PubMed Central

    Borkowski, Teresa A.; Letterio, John J.; Farr, Andrew G.; Udey, Mark C.

    1996-01-01

    Transforming growth factor β1 (TGF-β1) regulates leukocytes and epithelial cells. To determine whether the pleiotropic effects of TGF-β1, a cytokine that is produced by both keratinocytes and Langerhans cells (LC), extend to epidermal leukocytes, we characterized LC (the epidermal contingent of the dendritic cell [DC] lineage) and dendritic epidermal T cells (DETC) in TGF-β1 null (TGF-β1 −/−) mice. I-A+ LC were not detected in epidermal cell suspensions or epidermal sheets prepared from TGF-β1 −/− mice, and epidermal cell suspensions were devoid of allostimulatory activity. In contrast, TCR-γδ+ DETC were normal in number and appearance in TGF-β1 −/− mice and, importantly, DETC represented the only leukocytes in the epidermis. Immunolocalization studies revealed CD11c+ DC in lymph nodes from TGF-β1 −/− mice, although gp40+ DC were absent. Treatment of TGF-β1 −/− mice with rapamycin abrogated the characteristic inflammatory wasting syndrome and prolonged survival indefinitely, but did not result in population of the epidermis with LC. Thus, the LC abnormality in TGF-β1 −/− mice is not a consequence of inflammation in skin or other organs, and LC development is not simply delayed in these animals. We conclude that endogenous TGF-β1 is essential for normal murine LC development or epidermal localization. PMID:8976197

  16. Null Killing vectors

    NASA Astrophysics Data System (ADS)

    Lukács, B.; Perjés, Z.; Sebestyén, Á.

    1981-06-01

    Space-times admitting a null Killing vector are studied, using the Newman-Penrose spin coefficient formalism. The properties of the eigenrays (principal null curves of the Killing bivector) are shown to be related to the twist of the null Killing vector. Among the electrovacs, the ones containing a null Maxwell field turn out to belong to the twist-free class. An electrovac solution is obtained for which the null Killing vector is twisting and has geodesic and shear-free eigenrays. This solution is parameterless and appears to be the field of a zero-mass, spinning, and charged source.

  17. Increasing brain angiotensin converting enzyme 2 activity decreases anxiety-like behavior in male mice by activating central Mas receptors.

    PubMed

    Wang, Lei; de Kloet, Annette D; Pati, Dipanwita; Hiller, Helmut; Smith, Justin A; Pioquinto, David J; Ludin, Jacob A; Oh, S Paul; Katovich, Michael J; Frazier, Charles J; Raizada, Mohan K; Krause, Eric G

    2016-06-01

    Over-activation of the brain renin-angiotensin system (RAS) has been implicated in the etiology of anxiety disorders. Angiotensin converting enzyme 2 (ACE2) inhibits RAS activity by converting angiotensin-II, the effector peptide of RAS, to angiotensin-(1-7), which activates the Mas receptor (MasR). Whether increasing brain ACE2 activity reduces anxiety by stimulating central MasR is unknown. To test the hypothesis that increasing brain ACE2 activity reduces anxiety-like behavior via central MasR stimulation, we generated male mice overexpressing ACE2 (ACE2 KI mice) and wild type littermate controls (WT). ACE2 KI mice explored the open arms of the elevated plus maze (EPM) significantly more than WT, suggesting increasing ACE2 activity is anxiolytic. Central delivery of diminazene aceturate, an ACE2 activator, to C57BL/6 mice also reduced anxiety-like behavior in the EPM, but centrally administering ACE2 KI mice A-779, a MasR antagonist, abolished their anxiolytic phenotype, suggesting that ACE2 reduces anxiety-like behavior by activating central MasR. To identify the brain circuits mediating these effects, we measured Fos, a marker of neuronal activation, subsequent to EPM exposure and found that ACE2 KI mice had decreased Fos in the bed nucleus of stria terminalis but had increased Fos in the basolateral amygdala (BLA). Within the BLA, we determined that ∼62% of GABAergic neurons contained MasR mRNA and expression of MasR mRNA was upregulated by ACE2 overexpression, suggesting that ACE2 may influence GABA neurotransmission within the BLA via MasR activation. Indeed, ACE2 overexpression was associated with increased frequency of spontaneous inhibitory postsynaptic currents (indicative of presynaptic release of GABA) onto BLA pyramidal neurons and central infusion of A-779 eliminated this effect. Collectively, these results suggest that ACE2 may reduce anxiety-like behavior by activating central MasR that facilitate GABA release onto pyramidal neurons within the

  18. In Situ Aerosol-Size Distributions and Clear-Column Radiative Closure During ACE-2

    NASA Technical Reports Server (NTRS)

    Collins, R.; Jonsson, H. H.; Seinfeld, J. H.; Flagan, R. C.; Gasso, S.; Hegg, D. A.; Russell, P. B.; Schmid, B.; Livingston, J. M.; Oestroem, E.

    1999-01-01

    As part of the second Aerosol Characterization Experiment (ACE-2) during June and July of 1997, aerosol-size distributions were measured on board the CIRPAS Pelican aircraft through the use of a Differential Mobility Analyzer (DMA) and 2 Optical Particle Counters (OPCs). During the campaign, the boundary-layer aerosol typically possessed characteristics representative of a background marine aerosol or a continentally influenced aerosol, while the free-tropospheric aerosol was characterized by the presence or absence of a Saharan dust layer. A range of radiative closure comparisons were made using the data obtained during vertical profiles flown on 4 missions. Of particular interest here are the comparisons made between the optical properties as determined through the use of measured aerosol-size distributions and those measured directly by an airborne 14-wavelength sunphotometer and 3 nephelometers. Variations in the relative humidity associated with each of the direct measurements required consideration of the hygroscopic properties of the aerosol for size-distribution-based calculations. Simultaneous comparison with such a wide range of directly-measured optical parameters not only offers evidence of the validity of the physicochemical description of the aerosol when closure is achieved, but also provides insight into potential sources of error when some or all of the comparisons result in disagreement. Agreement between the derived and directly-measured optical properties varied for different measurements and for different cases. Averaged over the 4 case studies, the derived extinction coefficient at 525 nm exceeded that measured by the sunphotometer by 2.5% in the clean boundary layer, but underestimated measurements by 13% during pollution events. For measurements within the free troposphere, the mean derived extinction coefficient was 3.3% and 17% less than that measured by the sunphotometer during dusty and non-dusty conditions, respectively. Likewise

  19. In Situ Aerosol Size Distributions and Clear Column Radiative Closure During ACE-2

    NASA Technical Reports Server (NTRS)

    Collins, D. R.; Johnson, H. H.; Seinfeld, J. H.; Flagan, R. C.; Gasso, S.; Hegg, D. A.; Russell, P. B.; Schmid, B.; Livingston, J. M.; Oestroem, E.; Noone, K. J.; Russell, L. M.; Putaud, J. P.

    2000-01-01

    As part of the second Aerosol Characterization Experiment (ACE-2) during June and July of 1997, aerosol size distributions were measured on board the CIRPAS Pelican aircraft through the use of a DMA and two OPCS. During the campaign, the boundary layer aerosol typically possessed characteristics representative of a background marine aerosol or a continentally influenced aerosol, while the free tropospheric aerosol was characterized by the presence or absence of a Saharan dust layer. A range of radiative closure comparisons were made using the data obtained during vertical profiles flown on four missions. Of particular interest here are the comparisons made between the optical properties as determined through the use of measured aerosol size distributions and those measured directly by an airborne 14-wavelength sunphotometer and three nephelometers. Variations in the relative humidity associated with each of the direct measurements required consideration of the hygroscopic properties of the aerosol for size distribution based calculations. Simultaneous comparison with such a wide range of directly measured optical parameters not only offers evidence of the validity of the physicochemical description of the aerosol when closure is achieved, but also provides insight into potential sources of error when some or all of the comparisons result in disagreement. Agreement between the derived and directly measured optical properties varied for different measurements and for different cases. Averaged over the four case studies, the derived extinction coefficient at 525 nm exceeded that measured by the sunphotomoter by 2.5% in the clean boundary later, but underestimated measurements by 13% during pollution events. For measurements within the free troposphere, the mean derived extinction coefficient was 3.3% and 17% less than that measured by the sunphotometer during dusty and nondusty conditions, respectively. Likewise, averaged discrepancies between the derived and measured

  20. Impaired synthesis of erythropoietin, glutamine synthetase and metallothionein in the skin of NOD/SCID/gamma(c)(null) and Foxn1 nu/nu mice with misbalanced production of MHC class II complex.

    PubMed

    Danielyan, L; Verleysdonk, S; Buadze, M; Gleiter, C H; Buniatian, G H

    2010-06-01

    Most skin pathologies are characterized by unbalanced synthesis of major histocompatability complex II (MHC-II) proteins. Healthy skin keratinocytes simultaneously produce large amounts of MHC-II and regeneration-supporting proteins, e.g. erythropoietin (EPO), EPO receptor (EPOR), glutamine synthetase (GS) and metallothionein (MT). To investigate the level of regeneration-supporting proteins in the skin during misbalanced production of MHC-II, skin sections from nonobese diabetic/severe combined immunodeficient (NOD/SCID)/gamma (c) (null) and or Foxn1 nu/nu mice which are a priory known to under- and over-express MHC II, respectively, were used. Double immunofluorescence analysis of NOD/SCID/gamma (c) (null) skin sections showed striking decrease in expression of MHC-II, EPO, GS and MT. In Foxn1 nu/nu mouse skin, GS was strongly expressed in epidermis and in hair follicles (HF), which lacked EPO. In nude mouse skin EPO and MHC-II were over-expressed in dermal fibroblasts and they were completely absent from cortex, channel, medulla and keratinocytes surrounding the HF, suggest a role for EPO in health and pathology of hair follicle. The level of expression of EPO and GS in both mutant mice was confirmed by results of Western blot analyses. Strong immunoresponsiveness of EPOR in the hair channels of NOD/SCID/gamma (c) (null) mouse skin suggests increased requirements of skin cells for EPO and possible benefits of exogenous EPO application during disorders of immune system accompanied by loss MHC-II in skin cells.

  1. Clear Sky Column Closure Studies of Urban-Marine and Mineral-Dust Aerosols Using Aircraft, Ship, Satellite and Ground-Based Measurements in ACE-2

    NASA Technical Reports Server (NTRS)

    Schmid, Beat; Russell, Philip B.; Livingston, John M.; Gasso, Santiago; Hegg, Dean A.; Collins, Donald R.; Flagan, Richard C.; Seinfeld, John H.; Oestroem, Elisabeth; Noone, Kevin J.; Durkee, Philip A.; Jonsson, Haflidi H.; Welton, Ellsworth J.; Voss, Kenneth J.; Gordon, Howard R.; Formenti, Paola; Andreae, Meinrat O.; Kapustin, Vladimir N.; Bates, Timothy S.; Quinn, Patricia K.

    2000-01-01

    As part of the second Aerosol Characterization Experiment (ACE-2), European urban-marine and African mineral-dust aerosols were measured aboard the Pelican aircraft, the Research Vessel Vodyanitskiy from the ground and from satellites.

  2. ACE2 activation by xanthenone prevents leptin-induced increases in blood pressure and proteinuria during pregnancy in Sprague-Dawley rats.

    PubMed

    Ibrahim, Hisham Saleh; Froemming, Gabrielle Ruth Anisah; Omar, Effat; Singh, Harbindar Jeet

    2014-11-01

    This study investigates the effect of ACE2 activation on leptin-induced changes in systolic blood pressure (SBP), proteinuria, endothelial activation and ACE2 expression during pregnancy in Sprague-Dawley rats. Pregnant rats were given subcutaneous injection of either saline, or leptin, or leptin plus xanthenone (ACE2 activator), or xanthenone (XTN) alone. SBP, serum ACE, ACE2, endothelin-1, E-selectin and ICAM-1 levels were estimated; also their gene expressions were determined in the kidney and aorta respectively. Compared to control, SBP was higher in the leptin-only treated group (P<0.001) and lower in rats treated with xanthenone alone (P<0.01). Proteinuria, markers of endothelial activation were significantly higher than controls in leptin-only treated rats (P<0.05). ACE2 activity and expression were lower in leptin-only treated rats when compared to controls (P<0.05). It seems, leptin administration during pregnancy significantly increases SBP, proteinuria, endothelial activation, but decreases ACE2 level and expression. These effects are prevented by concurrent administration of xanthenone.

  3. The transcription factor Ace2 and its paralog Swi5 regulate ethanol production during static fermentation through their targets Cts1 and Rps4a in Saccharomyces cerevisiae.

    PubMed

    Wu, Yao; Du, Jie; Xu, Guoqiang; Jiang, Linghuo

    2016-05-01

    Saccharomyces cerevisiae is the most widely used fermentation organism for ethanol production. However, the gene expression regulatory networks behind the ethanol fermentation are still not fully understood. Using a static fermentation model, we examined the ethanol yields on biomass of deletion mutants for 77 yeast genes encoding nonessential transcription factors, and found that deletion mutants for ACE2 and SWI5 showed dramatically increased ethanol yields. Overexpression of ACE2 or SWI5 in wild type cells reduced their ethanol yields. Furthermore, among the 34 target genes regulated by Ace2 and Swi5, deletion of CTS1,RPS4a,SIC1,EGT2,DSE2, or SCP160 led to increased ethanol yields, with the former two showing higher effects. Overexpression of CTS1 or RPS4a in both ace2/ace2 and swi5/swi5 mutants reduced their ethanol yields. In contrast, deletion of MCR1 or HO significantly decreased ethanol yields, with the former one showing the highest effect. Therefore, Ace2 and Swi5 are two negative regulators of ethanol yield during static fermentation of yeast cells, and both CTS1 and RPS4a are major effectors mediating these two transcription factors in regulating ethanol production.

  4. Measurement and Analysis of Atmospheric Spectral Optical Depths with NASA Ames Airborne Sunphotometers During TARFOX and ACE-2

    NASA Technical Reports Server (NTRS)

    Livingston, John M.

    1997-01-01

    In accordance with the scope of work of this contract, the following tasks were undertaken and completed during the course of the contract: (1) Participation in the design and development of the 14-channel Ames Airborne Tracking Sunphotometer (AATS-14), including the development and implementation of Visual Basic software for real-time data processing and display and post-acquisition data reduction and analysis. (2) Operation of the six-channel Ames Airborne Tracking Sunphotometer (AATS-6) aboard the University of Washington C-131A during TARFOX and in-field analysis and presentation of data acquired with the AATS-6. (3) Post-mission analysis of data acquired during TARFOX with the AATS-6 and the AATS-14. (4) Pre-TARFOX calibration of the AATS-6 at Mauna Loa Observatory in May 1996, and post-TARFOX calibration of the AATS-6 and AATS- 14 at Zugspitze, Germany in October 1996, including analyses of all data sets. (5) Analysis of AATS-14 airborne calibration data acquired on 17 November 1996 during a late afternoon Pelican flight over the central California coast. (6) Operational training, instrument preparation, field coordination, and analysis of shipboard measurements of aerosol optical depth with the AATS-6 during ACE-2. (7) Coordination of data acquisition with the AATS-14 aboard the Pelican during ACE-2 and in-field preliminary data analysis and presentation. (8) Calibration of the AATS-6 and AATS-14 in April/May 1997 at Mauna Loa prior to ACE-2, and post-mission calibration of the AATS-6 at Mauna Loa in August 1997.

  5. Podocyte-specific overexpression of human angiotensin-converting enzyme 2 attenuates diabetic nephropathy in mice.

    PubMed

    Nadarajah, Renisha; Milagres, Rosangela; Dilauro, Marc; Gutsol, Alex; Xiao, Fengxia; Zimpelmann, Joseph; Kennedy, Chris; Wysocki, Jan; Batlle, Daniel; Burns, Kevin D

    2012-08-01

    Angiotensin-converting enzyme 2 (ACE2) degrades angiotensin II to angiotensin-(1-7) and is expressed in podocytes. Here we overexpressed ACE2 in podocytes in experimental diabetic nephropathy using transgenic methods where a nephrin promoter drove the expression of human ACE2. Glomeruli from these mice had significantly increased mRNA, protein, and activity of ACE2 compared to wild-type mice. Male mice were treated with streptozotocin to induce diabetes. After 16 weeks, there was no significant difference in plasma glucose levels between wild-type and transgenic diabetic mice. Urinary albumin was significantly increased in wild-type diabetic mice at 4 weeks, whereas albuminuria in transgenic diabetic mice did not differ from wild-type nondiabetic mice. However, this effect was transient and by 16 weeks both transgenic and nontransgenic diabetic mice had similar rates of proteinuria. Compared to wild-type diabetic mice, transgenic diabetic mice had an attenuated increase in mesangial area, decreased glomerular area, and a blunted decrease in nephrin expression. Podocyte numbers decreased in wild-type diabetic mice at 16 weeks, but were unaffected in transgenic diabetic mice. At 8 weeks, kidney cortical expression of transforming growth factor-β1 was significantly inhibited in transgenic diabetic mice as compared to wild-type diabetic mice. Thus, the podocyte-specific overexpression of human ACE2 transiently attenuates the development of diabetic nephropathy.

  6. Differential post-surgical metastasis and survival in SCID, NOD-SCID and NOD-SCID-IL-2Rγ(null) mice with parental and subline variants of human breast cancer: implications for host defense mechanisms regulating metastasis.

    PubMed

    Milsom, Chloe C; Lee, Christina R; Hackl, Christina; Man, Shan; Kerbel, Robert S

    2013-01-01

    We compare for the first time, the metastatic aggressiveness of the parental MDA-MB-231 breast cancer cell line and two luciferase-tagged in vivo-derived and selected pro-metastatic variants (LM2-4/luc⁺ and 164/8-1B/luc⁺ in SCID, NOD-SCID and NOD-SCID-IL-2Rγ(null) (NSG) mice following orthotopic implantation and primary tumour resection. The variants are known to be more aggressively metastatic in SCID mice, compared to the parental line which has limited spontaneous metastatic competence in these mice. When 2×10⁶ cells were injected into the mammary fat pad, the growth of the resultant primary tumours was identical for the various cell lines in the three strains of mice. However, metastatic spread of all three cell lines, including the MDA-MB-231 parental cell line, was strikingly more aggressive in the highly immunocompromised NSG mice compared to both NOD-SCID and SCID mice, resulting in extensive multi-organ metastases and a significant reduction in overall survival. While these studies were facilitated by monitoring post-surgical spontaneous metastases using whole body bioluminescence imaging, we observed that the luciferase-tagged parental line showed altered growth and diminished metastatic properties compared to its untagged counterpart. Our results are the first to show that host immunity can have a profound impact on the spread of spontaneous visceral metastases and survival following resection of a primary tumour in circumstances where the growth of primary tumours is not similarly affected; as such they highlight the importance of immunity in the metastatic process, and by extension, suggest certain therapeutic strategies that may have a significant impact on reducing metastasis.

  7. Differential Post-Surgical Metastasis and Survival in SCID, NOD-SCID and NOD-SCID-IL-2Rγnull Mice with Parental and Subline Variants of Human Breast Cancer: Implications for Host Defense Mechanisms Regulating Metastasis

    PubMed Central

    Milsom, Chloe C.; Lee, Christina R.; Hackl, Christina; Man, Shan; Kerbel, Robert S.

    2013-01-01

    We compare for the first time, the metastatic aggressiveness of the parental MDA-MB-231 breast cancer cell line and two luciferase-tagged in vivo-derived and selected pro-metastatic variants (LM2-4/luc+ and 164/8-1B/luc+) in SCID, NOD-SCID and NOD-SCID-IL-2Rγnull (NSG) mice following orthotopic implantation and primary tumour resection. The variants are known to be more aggressively metastatic in SCID mice, compared to the parental line which has limited spontaneous metastatic competence in these mice. When 2×106 cells were injected into the mammary fat pad, the growth of the resultant primary tumours was identical for the various cell lines in the three strains of mice. However, metastatic spread of all three cell lines, including the MDA-MB-231 parental cell line, was strikingly more aggressive in the highly immunocompromised NSG mice compared to both NOD-SCID and SCID mice, resulting in extensive multi-organ metastases and a significant reduction in overall survival. While these studies were facilitated by monitoring post-surgical spontaneous metastases using whole body bioluminescence imaging, we observed that the luciferase-tagged parental line showed altered growth and diminished metastatic properties compared to its untagged counterpart. Our results are the first to show that host immunity can have a profound impact on the spread of spontaneous visceral metastases and survival following resection of a primary tumour in circumstances where the growth of primary tumours is not similarly affected; as such they highlight the importance of immunity in the metastatic process, and by extension, suggest certain therapeutic strategies that may have a significant impact on reducing metastasis. PMID:23967178

  8. Chronic Proliferative Dermatitis in Sharpin Null Mice: Development of an Autoinflammatory Disease in the Absence of B and T Lymphocytes and IL4/IL13 Signaling

    PubMed Central

    Potter, Christopher S.; Wang, Zhe; Silva, Kathleen A.; Kennedy, Victoria E.; Stearns, Timothy M.; Burzenski, Lisa; Shultz, Leonard D.; HogenEsch, Harm; Sundberg, John P.

    2014-01-01

    SHARPIN is a key regulator of NFKB and integrin signaling. Mice lacking Sharpin develop a phenotype known as chronic proliferative dermatitis (CPDM), typified by progressive epidermal hyperplasia, apoptosis of keratinocytes, cutaneous and systemic eosinophilic inflammation, and hypoplasia of secondary lymphoid organs. Rag1−/− mice, which lack mature B and T cells, were crossed with Sharpin−/− mice to examine the role of lymphocytes in CDPM. Although inflammation in the lungs, liver, and joints was reduced in these double mutant mice, dermatitis was not reduced in the absence of functional lymphocytes, suggesting that lymphocytes are not primary drivers of the inflammation in the skin. Type 2 cytokine expression is increased in CPDM. In an attempt to reduce this aspect of the phenotype, Il4ra−/− mice, unresponsive to both IL4 and IL13, were crossed with Sharpin−/− mice. Double homozygous Sharpin−/−, Il4ra−/− mice developed an exacerbated granulocytic dermatitis, acute system inflammation, as well as hepatic necrosis and mineralization. High expression of CHI3L4, normally seen in CPDM skin, was abolished in Sharpin−/−, Il4ra−/− double mutant mice indicating the crucial role of IL4 and IL13 in the expression of this protein. Cutaneous eosinophilia persisted in Sharpin−/−, Il4ra−/− mice, although expression of Il5 mRNA was reduced and the expression of Ccl11 and Ccl24 was completely abolished. TSLP and IL33 were both increased in the skin of Sharpin−/− mice and this was maintained in Sharpin−/−, Il4ra−/− mice suggesting a role for TSLP and IL33 in the eosinophilic dermatitis in SHARPIN-deficient mice. These studies indicate that cutaneous inflammation in SHARPIN-deficient mice is autoinflammatory in nature developing independently of B and T lymphocytes, while the systemic inflammation seen in CPDM has a strong lymphocyte-dependent component. Both the cutaneous and systemic inflammation is enhanced by loss of IL4 and

  9. Airborne Sunphotometer, Airborne in-situ, Space-borne, and Ground-Based Measurements of Troposoheric Aerosol in Ace-2

    NASA Technical Reports Server (NTRS)

    Schmid, Beat; Collins, D.; Gasso, S.; Ostrom, E.; Powell, D.; Welton, E.; Durkee, P.; Livingstron, J.; Russell, P.; Flagan, R.; Hipskind, R. Stephen (Technical Monitor)

    2000-01-01

    We report on clear-sky column closure experiments performed in the Canary Islands during the second Aerosol Characterization Experiment (ACE-2) in June/July 1997. We present results obtained by combining airborne sunphotometer and in-situ aerosol measurements taken aboard the Pelican aircraft, space-borne NOAA/AVHRR data and ground-based lidars A wide range of aerosol types was encountered throughout the ACE-2 area, including background Atlantic marine, European pollution-derived, and African mineral dust. During !he two days discussed here, vertical profiles flown in cloud free air masses revealed three distinctly different layers: a marine boundary layer (MBL) with varying pollution levels, an elevated dust layer, and a very clean layer between the MBL and the dust layer. We found that the presence of the elevated dust layer removes the good agreement between satellite and sunphotometer AOD usually found in the absence of the dust layer. Using size-resolved composition information we have computed optical properties of the ambient aerosol from the in-situ measurements and subsequently compared those to the sunphotometer results. In the dust, the agreement in layer aerosol optical depth (380-1060 nm) is 3-8%. In the MBL there is tendency for the in-situ results to be slightly lower than the sunphotometer measurements (10-17% at 525 nm), but these differences are within the combined error bars of the measurements and computations.

  10. SARS coronavirus, but not human coronavirus NL63, utilizes cathepsin L to infect ACE2-expressing cells.

    PubMed

    Huang, I-Chueh; Bosch, Berend Jan; Li, Fang; Li, Wenhui; Lee, Kyoung Hoa; Ghiran, Sorina; Vasilieva, Natalya; Dermody, Terence S; Harrison, Stephen C; Dormitzer, Philip R; Farzan, Michael; Rottier, Peter J M; Choe, Hyeryun

    2006-02-10

    Viruses require specific cellular receptors to infect their target cells. Angiotensin-converting enzyme 2 (ACE2) is a cellular receptor for two divergent coronaviruses, SARS coronavirus (SARS-CoV) and human coronavirus NL63 (HCoV-NL63). In addition to hostcell receptors, lysosomal cysteine proteases are required for productive infection by some viruses. Here we show that SARS-CoV, but not HCoV-NL63, utilizes the enzymatic activity of the cysteine protease cathepsin L to infect ACE2-expressing cells. Inhibitors of cathepsin L blocked infection by SARS-CoV and by a retrovirus pseudotyped with the SARS-CoV spike (S) protein but not infection by HCoV-NL63 or a retrovirus pseudotyped with the HCoV-NL63 S protein. Expression of exogenous cathepsin L substantially enhanced infection mediated by the SARS-CoV S protein and by filovirus GP proteins but not by the HCoV-NL63 S protein or the vesicular stomatitis virus G protein. Finally, an inhibitor of endosomal acidification had substantially less effect on infection mediated by the HCoV-NL63 S protein than on that mediated by the SARS-CoV S protein. Our data indicate that two coronaviruses that utilize a common receptor nonetheless enter cells through distinct mechanisms.

  11. Tumor necrosis factor-alpha convertase (ADAM17) mediates regulated ectodomain shedding of the severe-acute respiratory syndrome-coronavirus (SARS-CoV) receptor, angiotensin-converting enzyme-2 (ACE2).

    PubMed

    Lambert, Daniel W; Yarski, Mike; Warner, Fiona J; Thornhill, Paul; Parkin, Edward T; Smith, A Ian; Hooper, Nigel M; Turner, Anthony J

    2005-08-26

    Angiotensin-converting enzyme-2 (ACE2) is a critical regulator of heart function and a cellular receptor for the causative agent of severe-acute respiratory syndrome (SARS), SARS-CoV (coronavirus). ACE2 is a type I transmembrane protein, with an extracellular N-terminal domain containing the active site and a short intracellular C-terminal tail. A soluble form of ACE2, lacking its cytosolic and transmembrane domains, has been shown to block binding of the SARS-CoV spike protein to its receptor. In this study, we examined the ability of ACE2 to undergo proteolytic shedding and investigated the mechanisms responsible for this shedding event. We demonstrated that ACE2, heterologously expressed in HEK293 cells and endogenously expressed in Huh7 cells, undergoes metalloproteinase-mediated, phorbol ester-inducible ectodomain shedding. By using inhibitors with differing potency toward different members of the ADAM (a disintegrin and metalloproteinase) family of proteases, we identified ADAM17 as a candidate mediator of stimulated ACE2 shedding. Furthermore, ablation of ADAM17 expression using specific small interfering RNA duplexes reduced regulated ACE2 shedding, whereas overexpression of ADAM17 significantly increased shedding. Taken together, these data provided direct evidence for the involvement of ADAM17 in the regulated ectodomain shedding of ACE2. The identification of ADAM17 as the protease responsible for ACE2 shedding may provide new insight into the physiological roles of ACE2.

  12. The application of hepatic P450 reductase null gpt delta mice in studying the role of hepatic P450 in genotoxic carcinogen 4-(methylnitrosamino)-1-(3-pyridyl)-1-butanone-induced mutagenesis.

    PubMed

    Luan, Yang; Xing, Guozhen; Qi, Xinming; Wu, Mengjun; Li, Chenggang; Yao, Jun; Gong, Likun; Nohmi, Takehiko; Gu, Jun; Zhou, Wanhong; Zheng, Saijing; Ren, Jin

    2012-11-01

    The cytochrome P450 (P450 or CYP) is involved in both detoxification and metabolic activation of many carcinogens. In order to identify the role of hepatic P450 in the mutagenesis of genotoxic carcinogens, we generated a novel hepatic P450 reductase null (HRN) gpt delta mouse model, which lacks functional hepatic P450 on a gpt delta mouse background. In this study, 4-(methylnitrosamino)-1-(3-pyridyl)-1-butanone (NNK) was used to treat HRN gpt delta mice and control littermates. Gene mutations in the liver and lungs were detected, and mutation spectra were analyzed. Pharmacokinetic analyses were performed, and tissue levels of NNK and metabolite were determined. NNK-induced mutant frequencies (MFs) were equivalent to spontaneous MFs in the liver, but increased more than 3 times in the lungs of HRN gpt delta mice compared to control mice. NNK-induced mutation spectra showed no difference between HRN gpt delta mice and control littermates. Toxicokinetic studies revealed reduced clearance of NNK with elevated tissue concentrations in HRN gpt delta mice. To our knowledge, these are the first data demonstrating that NNK cannot induce mutagenesis in the liver without P450 metabolic activation, but can induce mutagenesis in lungs by a hepatic P450-independent mechanism. Moreover, our data show that hepatic P450 plays a major role in the systemic clearance of NNK, thereby protecting the lungs against NNK-induced mutagenesis. Our model will be useful in establishing the role of hepatic versus extrahepatic P450-mediated mutagenesis, and the relative contributions of P450 compared to other biotransformation enzymes in the genotoxic carcinogens' activation.

  13. Inquiries into the Biological Significance of Transmembrane AMPA Receptor Regulatory Protein (TARP) γ-8 Through Investigations of TARP γ-8 Null Mice§.

    PubMed

    Gleason, Scott D; Kato, Akihiko; Bui, Hai H; Thompson, Linda K; Valli, Sabrina N; Stutz, Patrick V; Kuo, Ming-Shang; Falcone, Julie F; Anderson, Wesley H; Li, Xia; Witkin, Jeffrey M

    2015-01-01

    Transmembrane AMPA (α-amino-3-hydroxy-5-methyl-4-isoxazolepropionic acid) receptor regulatory protein (TARP) γ-8 is an auxiliary protein associated with some AMPA receptors. Most strikingly, AMPA receptors associated with this TARP have a relatively high localization in the hippocampus. TARP γ-8 also modifies the pharmacology and trafficking of AMPA receptors. However, to date there is little understanding of the biological significance of this auxiliary protein. In the present set of studies we provide a characterization of the differential pharmacology and behavioral consequences of deletion of TARP γ-8 by comparing the wild type (WT) and γ-8 -/- (knock-out, KO) mouse. KO mice were mildly hyperactive in a locomotor arena but not in other environments compared to WT mice. Additionally, the KO mice demonstrated enhanced locomotor stimulatory effects of both d-amphetamine and phencyclidine. Marble-burying and digging behaviors were dramatically reduced in KO mice. In another assay that can detect anxiety-like phenotypes, the elevated plus maze, no differences were observed in overall movement or open arm entries. In the forced-swim assay, KO mice displayed decreases in immobility time like the antidepressant imipramine and the AMPA receptor potentiator, LY392098. In KO mice, the antidepressant-like effects of LY392098 were prevented whereas the effects of imipramine were unaffected. Convulsions were induced by pentylenetetrazole, N-methyl-D-aspartate, and by kainic acid. However, in KO mice, kainic acid produced less tonic convulsions and lethality. KO mice had reduced levels of norepinephrine in hippocampus and cerebellum but not in hypothalamus or prefrontal cortex, decreased levels of cAMP in hippocampus, and increased levels of acetylcholine in the hypothalamus and prefrontal cortex. KO mice displayed decreased turnover of dopamine and increased histamine turnover in multiple brain areas In contrast, serotonin and its metabolites were not significantly

  14. Direct Comparison of Mice Null for Liver or Intestinal Fatty Acid-binding Proteins Reveals Highly Divergent Phenotypic Responses to High Fat Feeding*

    PubMed Central

    Gajda, Angela M.; Zhou, Yin Xiu; Agellon, Luis B.; Fried, Susan K.; Kodukula, Sarala; Fortson, Walter; Patel, Khamoshi; Storch, Judith

    2013-01-01

    The enterocyte expresses two fatty acid-binding proteins (FABP), intestinal FABP (IFABP; FABP2) and liver FABP (LFABP; FABP1). LFABP is also expressed in liver. Despite ligand transport and binding differences, it has remained uncertain whether these intestinally coexpressed proteins, which both bind long chain fatty acids (FA), are functionally distinct. Here, we directly compared IFABP−/− and LFABP−/− mice fed high fat diets containing long chain saturated or unsaturated fatty acids, reasoning that providing an abundance of dietary lipid would reveal unique functional properties. The results showed that mucosal lipid metabolism was indeed differentially modified, with significant decreases in FA incorporation into triacylglycerol (TG) relative to phospholipid (PL) in IFABP−/− mice, whereas LFABP−/− mice had reduced monoacylglycerol incorporation in TG relative to PL, as well as reduced FA oxidation. Interestingly, striking differences were found in whole body energy homeostasis; LFABP−/− mice fed high fat diets became obese relative to WT, whereas IFABP−/− mice displayed an opposite, lean phenotype. Fuel utilization followed adiposity, with LFABP−/− mice preferentially utilizing lipids, and IFABP−/− mice preferentially metabolizing carbohydrate for energy production. Changes in body weight and fat may arise, in part, from altered food intake; mucosal levels of the endocannabinoids 2-arachidonoylglycerol and arachidonoylethanolamine were elevated in LFABP−/−, perhaps contributing to increased energy intake. This direct comparison provides evidence that LFABP and IFABP have distinct roles in intestinal lipid metabolism; differential intracellular functions in intestine and in liver, for LFABP−/− mice, result in divergent downstream effects at the systemic level. PMID:23990461

  15. Environmental enrichment enhances neurogranin expression and hippocampal learning and memory but fails to rescue the impairments of neurogranin null mutant mice.

    PubMed

    Huang, Freesia L; Huang, Kuo-Ping; Wu, Junfang; Boucheron, Catherine

    2006-06-07

    Environmental enrichment is known to enhance hippocampal neurogenesis and cognitive functions. Neurogranin (Ng), a specific substrate of protein kinase C (PKC), is abundantly expressed in brain regions important for cognitive functions. Deletion of Ng in mice causes severe deficits in spatial learning and long-term potentiation (LTP) in the hippocampal CA1 region. These Ng-/- mice, as compared with Ng+/+, respond poorly after treatment of their hippocampal slices with agents that activate signaling molecules important for learning and memory, including Ca2+/calmodulin-dependent protein kinase II (alphaCaMKII), PKC, protein kinase A (PKA), extracellular signal-regulated kinase (ERK), and cAMP response element-binding protein (CREB). In the present study, adult mice were housed in either regular home cages (control group) or more spacious cages with an exercise wheel and change of toys twice per week (enriched group) for at least 3 weeks. Enriched Ng+/+ and Ng+/- mice showed enhanced LTP in the hippocampal CA1 after high-frequency stimulation, but Ng-/- mice were affected only minimally. Behaviorally, the enriched Ng+/+ and Ng+/-, but not Ng-/- mice, performed significantly better than their respective control cohorts in Morris water maze and in step-down fear conditioning. Enriched Ng+/- mice also showed improvement in the radial arm maze. Quantitative immunoblot analyses showed that the enriched groups of all three genotypes exhibited elevated hippocampal levels of alphaCaMKII and CREB, but not ERK. Interestingly, enrichment caused a significant increase in hippocampal Ng levels both in Ng+/+ and Ng+/- mice that seemed to contribute to their improved LTP and behavioral performances. These results suggest that Ng gates the neuronal signaling reactions involved in learning and memory. During environmental enrichment, these Ng-regulated reactions are also critical for the enhancement of synaptic plasticity and cognitive functions.

  16. Mutations in Acetylcholinesterase2 (ace2) increase the insensitivity of acetylcholinesterase to fosthiazate in the root-knot nematode Meloidogyne incognita.

    PubMed

    Huang, Wen-Kun; Wu, Qin-Song; Peng, Huan; Kong, Ling-An; Liu, Shi-Ming; Yin, Hua-Qun; Cui, Ru-Qiang; Zhan, Li-Ping; Cui, Jiang-Kuan; Peng, De-Liang

    2016-11-29

    The root-knot nematode Meloidogyne incognita causes severe damage to continuously cropping vegetables. The control of this nematode relies heavily on organophosphate nematicides in China. Here, we described resistance to the organophosphate nematicide fosthiazate in a greenhouse-collected resistant population (RP) and a laboratory susceptible population (SP) of M. incognita. Fosthiazate was 2.74-fold less toxic to nematodes from RP than that from SP. Quantitative real-time PCR revealed that the acetylcholinesterase2 (ace2) transcription level in the RP was significantly higher than that in the SP. Eighteen nonsynonymous amino acid differences in ace2 were observed between the cDNA fragments of the RP and SP. The acetylcholinesterase (AChE) protein activity in the RP was significantly reduced compared with that in the SP. After knocking down the ace2 gene, the ace2 transcription level was significantly decreased, but no negative impact on the infection of juveniles was observed. The 50% lethal concentration of the RNAi RP population decreased 40%, but the inhibition rate of fosthiazate against AChE activity was significantly increased in RP population. Thus, the increased fosthiazate insensitivity in the M. incognita resistant population was strongly associated with mutations in ace2. These results provide valuable insights into the resistance mechanism of root-knot nematode to organophosphate nematicides.

  17. Mutations in Acetylcholinesterase2 (ace2) increase the insensitivity of acetylcholinesterase to fosthiazate in the root-knot nematode Meloidogyne incognita

    PubMed Central

    Huang, Wen-Kun; Wu, Qin-Song; Peng, Huan; Kong, Ling-An; Liu, Shi-Ming; Yin, Hua-Qun; Cui, Ru-Qiang; Zhan, Li-Ping; Cui, Jiang-Kuan; Peng, De-Liang

    2016-01-01

    The root-knot nematode Meloidogyne incognita causes severe damage to continuously cropping vegetables. The control of this nematode relies heavily on organophosphate nematicides in China. Here, we described resistance to the organophosphate nematicide fosthiazate in a greenhouse-collected resistant population (RP) and a laboratory susceptible population (SP) of M. incognita. Fosthiazate was 2.74-fold less toxic to nematodes from RP than that from SP. Quantitative real-time PCR revealed that the acetylcholinesterase2 (ace2) transcription level in the RP was significantly higher than that in the SP. Eighteen nonsynonymous amino acid differences in ace2 were observed between the cDNA fragments of the RP and SP. The acetylcholinesterase (AChE) protein activity in the RP was significantly reduced compared with that in the SP. After knocking down the ace2 gene, the ace2 transcription level was significantly decreased, but no negative impact on the infection of juveniles was observed. The 50% lethal concentration of the RNAi RP population decreased 40%, but the inhibition rate of fosthiazate against AChE activity was significantly increased in RP population. Thus, the increased fosthiazate insensitivity in the M. incognita resistant population was strongly associated with mutations in ace2. These results provide valuable insights into the resistance mechanism of root-knot nematode to organophosphate nematicides. PMID:27897265

  18. [Effect of Astragali Radix in improving early renal damage in metabolic syndrome rats through ACE2/Mas pathway].

    PubMed

    Wang, Qiong-ying; Liang, Wei; Jiang, Cheng; Li, Ning-yin; Xu, Han; Yang, Mi-na; Lin, Xin; Yu, Heng; Chang, Peng; Yu, Jing

    2015-11-01

    To study the expression of angiotensin converting enzyme 2 (ACE2) and angiotensin (Ang) 1-7 specific receptor Mas protain in renal blood vessels of metabolic syndrome ( MS) rats and its anti-oxidative effect. A total of 80 male SD rats were divided into four groups: the normal control group (NC, the same volume of normal saline), the MS group (high fat diet), the MS + Astragali Radix group (MS + HQ, 6 g x kg(-1) x d(-1) in gavage) and the MS + Valsartan group (MS + XST, 30 mg x kg(-1) x d(-1) in gavage). After four weeks of intervention, their general indexes, biochemical indexes and blood pressure were measured; plasma and renal tissue Ang II, malondialdehyde (MDA) and superoxide demutase (SOD) levels were measured with radioimmunoassay. The protein expressions of Mas receptor, AT1R, ACE and ACE2 were detected by western blot analysis. According to the result, compared with the NC group, the MS group and the MS + HQ group showed significant increases in systolic and diastolic pressures, body weight, fasting glucose, fasting insulin, triglycerides, free fatty acid and Ang II level of MS rats (P < 0.05). The MS + XST group showed notable decreases in systolic and diastolic pressures than that of the MS group. The MS group showed significant increases in the SOD activity and NO level and decrease in the MDA level after being intervened with Astragali Radix. ACE and AT1R protein expressions in renal tissues of the MS group were higher than that in the NC group, but with lower ACE2 and -Mas receptor expressions (all P < 0.05). Compared with the MS group, the MS + HQ group showed significant increase in Mas receptor expression in renal tissues, whereas the MS + XST group showed notable decrease in AT1R (all P < 0.05). In conclusion, Astragali Radix can increase the Mas receptor expressions in renal tissues, decrease ACE expression and change local Ang II, MDA, NO and SOD in kidneys, so as to protect early damages in renal tissues.

  19. Nulling Breadboard for DARWIN

    NASA Astrophysics Data System (ADS)

    Flatscher, Reinhold; Johann, Ulrich; Sodnik, Zoran

    2003-02-01

    This work is funded by ESA under ESTEC/Contract No. 14827/00/NL/CK. Astrium Germany has been awarded this first ESA breadboarding towards nulling interferometry. Interferometric nulling devices are essential ingredients in the TPF and DARWIN missions for suppressing the star light by a factor of 106 over a wide wavelength range in the mid infrared. The current DARWIN baseline concept comprises six telescopes. The coherent combination scheme in the nulling mode operation foresees three nulling assemblies in parallel. The breadboard serves to demonstrate the deep and stable null required for an operational instrument. The demonstrator operates in the near infrared to save costs but its principle is fully applicable to the mid infrared. The nulling device is based on an autobalancing Sagnac core offering just one critical beam combiner. Two different ways of achieving the required π phase shift are implemented: a) arbitrary phase shift by dispersive phase shifter plates b) phase shift of π using periscopes (image flip) The target simulator features two point sources of adjustable radiometry and angular separation, representing a strong star and a weak planet. In addition, the sources can be also used to simulate a double star for demonstrating the basic DARWIN imaging mode. The simulator can be operated in two styles, namely as wavefront dividing star/planet source and, alternatively, as an amplitude dividing source, providing highly symmetric wavefronts to both interferometer arms. Because of its representativity for the DARWIN situation, the latter mode is the preferred simulator for quantitative nulling experiments. The breadboard design has been finalized in January 2002 and verified by detailed simulations. The entire hardware has been manufactured by end of July. Currently, nulling and imaging measurements are in progress to validate the per-formance of the selected approach. The project is part of ESA´s technology preparatory program for DARWIN, paving the

  20. Residual Chemosensory Capabilities in Double P2X2/P2X3 Purinergic Receptor Null Mice: Intraoral or Postingestive Detection?

    PubMed Central

    Hallock, Robert M.; Tatangelo, Marco; Barrows, Jennell

    2009-01-01

    Mice lacking the purinergic receptors, P2X2 and P2X3 (P2X2/P2X3Dbl−/−), exhibit essentially no tastant-evoked activity in the chorda tympani and glossopharyngeal nerves and substantial loss of tastant-evoked behavior as measured in long-term intake experiments. To assess whether the residual chemically driven behaviors in these P2X2/P2X3Dbl−/− mice were attributable to postingestive detection or oropharyngeal detection of the compounds, we used brief access lickometer tests to assess the behavioral capabilities of the P2X2/P2X3Dbl−/− animals. The P2X2/P2X3Dbl−/− mice showed avoidance to high levels (10 mM quinine and 10–30 mM denatonium benzoate) of classical “bitter”-tasting stimuli in 24-h, 2-bottle preference tests but minimal avoidance of these substances in the lickometer tests, suggesting that the strong avoidance in the intake tests was largely mediated by post-oral chemosensors. Similarly, increases in consumption of 1 M sucrose by P2X2/P2X3Dbl−/− mice in long-term intake tests were not mirrored by increases in consumption of sucrose in lickometer tests, suggesting that sucrose detection in these mice is mediated by postingestive consequences. In contrast, in brief access tests, P2X2/P2X3Dbl−/− mice avoided citric acid and hydrochloric acid at the same concentrations as their wild-type counterparts, indicating that these weak acids activate oropharyngeal chemoreceptors. PMID:19833662

  1. Functional and molecular evidence for expression of the renin angiotensin system and ADAM17-mediated ACE2 shedding in COS7 cells

    PubMed Central

    Grobe, Nadja; Di Fulvio, Mauricio; Kashkari, Nada; Chodavarapu, Harshita; Somineni, Hari K.; Singh, Richa

    2015-01-01

    The renin angiotensin system (RAS) plays a vital role in the regulation of the cardiovascular and renal functions. COS7 is a robust and easily transfectable cell line derived from the kidney of the African green monkey, Cercopithecus aethiops. The aims of this study were to 1) demonstrate the presence of an endogenous and functional RAS in COS7, and 2) investigate the role of a disintegrin and metalloproteinase-17 (ADAM17) in the ectodomain shedding of angiotensin converting enzyme-2 (ACE2). Reverse transcription coupled to gene-specific polymerase chain reaction demonstrated expression of ACE, ACE2, angiotensin II type 1 receptor (AT1R), and renin at the transcript levels in total RNA cell extracts. Western blot and immunohistochemistry identified ACE (60 kDa), ACE2 (75 kDa), AT1R (43 kDa), renin (41 kDa), and ADAM17 (130 kDa) in COS7. At the functional level, a sensitive and selective mass spectrometric approach detected endogenous renin, ACE, and ACE2 activities. ANG-(1–7) formation (m/z 899) from the natural substrate ANG II (m/z 1,046) was detected in lysates and media. COS7 cells stably expressing shRNA constructs directed against endogenous ADAM17 showed reduced ACE2 shedding into the media. This is the first study demonstrating endogenous expression of the RAS and ADAM17 in the widely used COS7 cell line and its utility to study ectodomain shedding of ACE2 mediated by ADAM17 in vitro. The transfectable nature of this cell line makes it an attractive cell model for studying the molecular, functional, and pharmacological properties of the renal RAS. PMID:25740155

  2. Downregulation of ACE2/Ang-(1-7)/Mas axis promotes breast cancer metastasis by enhancing store-operated calcium entry.

    PubMed

    Yu, Changhui; Tang, Wei; Wang, Yuhao; Shen, Qiang; Wang, Bin; Cai, Chunqing; Meng, Xiaojing; Zou, Fei

    2016-07-01

    The renin-angiotensin system (RAS) is an important component of the tumor microenvironment and plays a key role in promoting cancer cell proliferation, angiogenesis, metabolism, migration and invasion. Meanwhile, the arm of angiotensin-converting enzyme (ACE)2/angiotensin-(1-7) [Ang-(1-7)]/Mas axis in connection with RAS is associated with anti-proliferative, vasodilatory and anti-metastatic properties. Previous studies have shown that Ang-(1-7) reduces the proliferation of orthotopic human breast tumor growth by inhibiting cancer-associated fibroblasts. However, the role of ACE/Ang-(1-7)/Mas axis in the metastasis of breast cancer cells is still unknown. In the present study, we found that ACE2 protein level is negatively correlated with the metastatic ability of breast cancer cells and breast tumor grade. Upregulation of ACE2/Ang-(1-7)/Mas axis inhibits breast cancer cell migration and invasion in vivo and in vitro. Mechanistically, ACE2/Ang-(1-7)/Mas axis activation inhibits store-operated calcium entry (SOCE) and PAK1/NF-κB/Snail1 pathways, and induces E-cadherin expression. In summary, our results demonstrate that downregulation of ACE2/Ang-(1-7)/Mas axis stimulates breast cancer metastasis through the activation of SOCE and PAK1/NF-κB/Snail1 pathways. These results provide new mechanisms by which breast cancer develop metastasis and shed light on developing novel anti-metastasis therapeutics for metastatic breast cancer by modulating ACE2/Ang-(1-7)/Mas axis.

  3. North Atlantic Aerosol Radiative Impacts Based on Satellite Measurements and Aerosol Intensive Properties from TARFOX and ACE-2

    NASA Technical Reports Server (NTRS)

    Russell, P. B.; Bergstrom, Robert W.; Schmid, B.; Livingston, J. M.

    2000-01-01

    We estimate the impact of North Atlantic aerosols on the net short-wave flux at the tropopause by combining satellite-derived aerosol optical depth (AOD) maps with model aerosol properties determined via closure analyses in TARFOX and ACE 2. We exclude African dust, primarily by restricting latitudes to 25-60 N. The analyses use in situ aerosol composition measurements and air- and ship-borne sun-photometer measurements of AOD spectra. The aerosol model yields computed flux sensitivities (dFlux/dAOD) that agree with measurements by airborne flux radiometers in TARFOX. Its midvisible single-scattering albedo is 0.9. which is in the range obtained from in situ measurements of scattering and absorption in both TARFOX and ACE 2. Combining satellite-derived AOD maps with the aerosol model yields maps of 24-hour average net radiative flux changes. For simultaneous AVHRR, radiance measurements exceeded the sunphotometer AODs by about 0.04. However. shipboard sunphotometer and AVHRR AODs agreed Within 0.02 for data acquired during satellite overflights on two other days. We discuss attempts to demonstrate column closure within the MBL by comparing shipboard sunphotometer AODs and values calculated from simultaneous shipboard in-situ aerosol size distribution measurements. These comparisons were mostly unsuccessful, but they illustrate the difficulties inherent in this type of closure analysis. Specifically, AODs derived from near-surface in-situ size distribution measurements are extremely sensitive to the assumed hygroscopic growth model that itself requires an assumption of particle composition as a function of height and size, to the radiosonde-measured relative humidity, and to the vertical profile of particle number. We investigate further the effects of hygroscopic particle growth within the MBL by using shipboard lidar aerosol backscatter profiles together with the sunphotometer AOD.

  4. Lung mechanics in the TIMP3 null mouse and its response to mechanical ventilation.

    PubMed

    Martin, Erica L; Truscott, Emily A; Bailey, Timothy C; Leco, Kevin J; McCaig, Lynda A; Lewis, James F; Veldhuizen, Ruud A W

    2007-03-01

    Tissue inhibitor of metalloproteinase-3 (TIMP3) null mice develop emphysema-like airspace enlargement due to an enzymatic imbalance. This study investigates how these abnormalities alter lung mechanics and the response to 2 different mechanical ventilation strategies. Phenotypically, TIMP3 null mice had increased compliance, and decreased resistance, tissue damping, and tissue elastance over wild-type controls. Decreased compliance and increased resistance were observed following the injurious ventilation strategy; however, the TIMP3 null response to both ventilation strategies was similar to wild-type mice. In conclusion, TIMP3 null mice have significant alterations in lung mechanics; however, this does not affect their response to ventilation.

  5. THE EMBRYOLETHALITY OF LIPOPOLYSACCHARIDE IN CD-1 AND METALLOTHIONEIN I-II NULL MICE: LACK OF A ROLE FOR INDUCED ZINC DEFICIENCY OR METALLOTHIONEIN INDUCTION

    EPA Science Inventory

    ABSTRACT

    Lipopolysaccharide (LPS) is embryolethal in CD-1 mice. LPS induces metallothionein (MT) via cytokines, including TNF-, IL-1 and IL-6, which initiate and maintain the acute phase response. Maternal hepatic MT induction in pregnant rats, by diverse toxicants, can ...

  6. Sex Differences in Ethanol's Anxiolytic Effect and Chronic Ethanol Withdrawal Severity in Mice with a Null Mutation of the 5α-Reductase Type 1 Gene.

    PubMed

    Tanchuck-Nipper, Michelle A; Ford, Matthew M; Hertzberg, Anna; Beadles-Bohling, Amy; Cozzoli, Debra K; Finn, Deborah A

    2015-05-01

    Manipulation of endogenous levels of the GABAergic neurosteroid allopregnanolone alters sensitivity to some effects of ethanol. Chronic ethanol withdrawal decreases activity and expression of 5α-reductase-1, an important enzyme in allopregnanolone biosynthesis encoded by the 5α-reductase-1 gene (Srd5a1). The present studies examined the impact of Srd5a1 deletion in male and female mice on several acute effects of ethanol and on chronic ethanol withdrawal severity. Genotype and sex did not differentially alter ethanol-induced hypothermia, ataxia, hypnosis, or metabolism, but ethanol withdrawal was significantly lower in female versus male mice. On the elevated plus maze, deletion of the Srd5a1 gene significantly decreased ethanol's effect on total entries versus wildtype (WT) mice and significantly decreased ethanol's anxiolytic effect in female knockout (KO) versus WT mice. The limited sex differences in the ability of Srd5a1 genotype to modulate select ethanol effects may reflect an interaction between developmental compensations to deletion of the Srd5a1 gene with sex hormones and levels of endogenous neurosteroids.

  7. Corneal haze phenotype in Aldh3a1-null mice: In vivo confocal microscopy and tissue imaging mass spectrometry.

    PubMed

    Chen, Ying; Jester, James V; Anderson, David M; Marchitti, Satori A; Schey, Kevin L; Thompson, David C; Vasiliou, Vasilis

    2016-12-27

    ALDH3A1 is a corneal crystallin that protects ocular tissues from ultraviolet radiation through catalytic and non-catalytic functions. In addition, ALDH3A1 plays a functional role in corneal epithelial homeostasis by simultaneously modulating proliferation and differentiation. We have previously shown that Aldh3a1 knockout mice in a C57B6/129sV mixed genetic background develop lens cataracts. In the current study, we evaluated the corneal phenotype of Aldh3a1 knockout mice bred into a C57B/6J congenic background (KO). In vivo confocal microscopy examination of KO and wild-type (WT) corneas revealed KO mice to exhibit corneal haze, manifesting marked light scattering from corneal stroma. This corneal phenotype was further characterized by Imaging Mass Spectrometry (IMS) with spatial resolution that revealed a trilayer structure based on differential lipid localization. In these preliminary studies, no differences were observed in lipid profiles from KO relative to WT mice; however, changes in protein profiles of acyl-CoA binding protein (m/z 9966) and histone H4.4 (m/z 11308) were found to be increased in the corneal epithelial layer of KO mice. This is the first study to use IMS to characterize endogenous proteins and lipids in corneal tissue and to molecularly explore the corneal haze phenotype. Taken together, the current study presents the first genetic animal model of cellular-induced corneal haze due to the loss of a corneal crystallin, and strongly supports the notion that ALDH3A1 is critical to cellular transparency. Finally, IMS represents a valuable new approach to reveal molecular changes underlying corneal disease.

  8. Augmented atherogenesis in ApoE-null mice co-exposed to polychlorinated biphenyls and 2,3,7,8-tetrachlorodibenzo-p-dioxin

    SciTech Connect

    Shan, Qiuli; Wang, Jing; Huang, Fengchen; Lv, Xiaowen; Ma, Min; Du, Yuguo

    2014-04-15

    2,3,7,8-Tetrachlorodibenzo-p-dioxin (TCDD) and polychlorinated biphenyls (PCBs) are persistent organic pollutants found as complex mixtures in the environment throughout the world. Therefore, humans are ubiquitously and simultaneously exposed to TCDD and PCBs. TCDD and PCBs alone have been linked to atherosclerosis. However, the effects of interactions or synergism between TCDD and PCBs on atherogenesis are unknown. We investigated the possible enhanced atherogenesis by co-exposure to TCDD and PCBs and the potential mechanism(s) involved in this enhancement. Male ApoE{sup −/−} mice were exposed to TCDD (15 μg/kg) and Aroclor1254 (55 mg/kg, a representative mixture of PCBs) alone or in combination by intraperitoneal injection four times over six weeks of duration. Our results showed that mice exposed to TCDD alone, but not Aroclor1254 alone, developed atherosclerotic lesions. Moreover, we found that atherosclerotic disease was exacerbated to the greatest extent in mice co-exposed to TCDD and Aroclor1254. The enhanced lesions correlated with several pro-atherogenic changes, including a marked increase in the accumulation of the platelet-derived chemokine PF4, and the expression of the proinflammatory cytokine MCP-1 and the critical immunity gene-RIG-I. Our data demonstrated that co-exposure to TCDD and Aroclor1254 markedly enhanced atherogenesis in ApoE{sup −/−} mice. Significantly, our observations suggest that combined exposure to TCDD and PCBs may be a greater cardiovascular health risk than previously anticipated from individual studies. - Highlights: • Augmented atherogenesis was found in ApoE{sup −/−} mice co-exposed to Aroclor1254 and TCDD. • Enhanced expression of PF4, MCP-1 and RIG-I correlated with augmented lesions. • POPs combination may be a greater cardiovascular health risk than individual POPs.

  9. The antidepressant-like effects of glutamatergic drugs ketamine and AMPA receptor potentiator LY 451646 are preserved in bdnf⁺/⁻ heterozygous null mice.

    PubMed

    Lindholm, Jesse S O; Autio, Henri; Vesa, Liisa; Antila, Hanna; Lindemann, Lothar; Hoener, Marius C; Skolnick, Phil; Rantamäki, Tomi; Castrén, Eero

    2012-01-01

    Accumulating evidence suggests that biogenic amine-based antidepressants act, at least in part, via regulation of brain-derived neurotrophic factor (BDNF) signaling. Biogenic amine-based antidepressants increase BDNF synthesis and activate its signaling pathway through TrkB receptors. Moreover, the antidepressant-like effects of these molecules are abolished in BDNF deficient mice. Glutamate-based drugs, including the NMDA antagonist ketamine, and the AMPA receptor potentiator LY 451646, mimic the effects of antidepressants in preclinical tests with high predictive validity. In humans, a single intravenous dose of ketamine produces an antidepressant effect that is rapid, robust and persistent. In this study, we examined the role of BDNF in expression of the antidepressant-like effects of ketamine and an AMPA receptor potentiator (LY 451646) in the forced swim test (FST). Ketamine and LY 451646 produced antidepressant-like effects in the FST in mice at 45 min after a single injection, but no effects were observed one week after a single ketamine injection. As previously reported, the effects of imipramine in the forced swim test were blunted in heterozygous BDNF knockout (bdnf(+/-)) mice. However ketamine and LY 451646 produced similar antidepressant-like responses in wildtype and bdnf(+/-) mice. Neither ketamine nor LY 451646 significantly influenced the levels BDNF or TrkB phosphorylation in the hippocampus when assessed at 45 min or 7 days after the drug administration. These data demonstrate that under the conditions tested, neither ketamine nor the AMPA-potentiator LY 451656 activate BDNF signaling, but produce a characteristic antidepressant-like response in heterozygous bdnf(+/-) mice. These data indicate that unlike biogenic amine-based agents, BDNF signaling does not play a pivotal role in the antidepressant effects of glutamate-based compounds. This article is part of a Special Issue entitled 'Anxiety and Depression'.

  10. Network Models Predict that Reduced Excitatory Fluctuations Can Give Rise to Hippocampal Network Hyper-Excitability in MeCP2-Null Mice

    PubMed Central

    Ho, Ernest C. Y.; Eubanks, James H.; Zhang, Liang; Skinner, Frances K.

    2014-01-01

    Rett syndrome is a severe pediatric neurological disorder caused by loss of function mutations within the gene encoding methyl CpG-binding protein 2 (MeCP2). Although MeCP2 is expressed near ubiquitously, the primary pathophysiology of Rett syndrome stems from impairments of nervous system function. One alteration within different regions of the MeCP2-deficient brain is the presence of hyper-excitable network responses. In the hippocampus, such responses exist despite there being an overall decrease in spontaneous excitatory drive within the network. In this study, we generated and used mathematical, neuronal network models to resolve this apparent paradox. We did this by taking advantage of previous mathematical modelling insights that indicated that decreased excitatory fluctuations, but not mean excitatory drive, more critically explain observed changes in hippocampal network oscillations from MeCP2-null mouse slices. Importantly, reduced excitatory fluctuations could also bring about hyper-excitable responses in our network models. Therefore, these results indicate that diminished excitatory fluctuations may be responsible for the hyper-excitable state of MeCP2-deficient hippocampal circuitry. PMID:24642514

  11. Exacerbation of experimental autoimmune encephalomyelitis in prion protein (PrPc)-null mice: evidence for a critical role of the central nervous system

    PubMed Central

    2012-01-01

    Background The cellular prion protein (PrPc) is a host-encoded glycoprotein whose transconformation into PrP scrapie (PrPSc) initiates prion diseases. The role of PrPc in health is still obscure, but many candidate functions have been attributed to the protein, both in the immune and the nervous systems. Recent data show that experimental autoimmune encephalomyelitis (EAE) is worsened in mice lacking PrPc. Disease exacerbation has been attributed to T cells that would differentiate into more aggressive effectors when deprived of PrPc. However, alternative interpretations such as reduced resistance of neurons to autoimmune insult and exacerbated gliosis leading to neuronal deficits were not considered. Method To better discriminate the contribution of immune cells versus neural cells, reciprocal bone marrow chimeras with differential expression of PrPc in the lymphoid or in the central nervous system (CNS) were generated. Mice were subsequently challenged with MOG35-55 peptide and clinical disease as well as histopathology were compared in both groups. Furthermore, to test directly the T cell hypothesis, we compared the encephalitogenicity of adoptively transferred PrPc-deficient versus PrPc-sufficient, anti-MOG T cells. Results First, EAE exacerbation in PrPc-deficient mice was confirmed. Irradiation exacerbated EAE in all the chimeras and controls, but disease was more severe in mice with a PrPc-deleted CNS and a normal immune system than in the reciprocal construction. Moreover, there was no indication that anti-MOG responses were different in PrPc-sufficient and PrPc-deficient mice. Paradoxically, PrPc-deficient anti-MOG 2D2 T cells were less pathogenic than PrPc-expressing 2D2 T cells. Conclusions In view of the present data, it can be concluded that the origin of EAE exacerbation in PrPc-ablated mice resides in the absence of the prion protein in the CNS. Furthermore, the absence of PrPc on both neural and immune cells does not synergize for disease worsening

  12. Ex vivo expanded human regulatory T cells delay islet allograft rejection via inhibiting islet-derived monocyte chemoattractant protein-1 production in CD34+ stem cells-reconstituted NOD-scid IL2rγnull mice.

    PubMed

    Xiao, Fang; Ma, Liang; Zhao, Min; Huang, Guocai; Mirenda, Vincenzo; Dorling, Anthony; Lechler, Robert; Lombardi, Giovanna

    2014-01-01

    Type 1 diabetes mellitus (T1DM) is an autoimmune disease caused by immune-mediated destruction of insulin-secreting β cells of the pancreas. Near complete dependence on exogenous insulin makes T1DM very difficult to control, with the result that patients are exposed to high blood glucose and risk of diabetic complications and/or intermittent low blood glucose that can cause unconsciousness, fits and even death. Allograft transplantation of pancreatic islets restores normoglycemia with a low risk of surgical complications. However, although successful immediately after transplantation, islets are progressively lost, with most of the patients requiring exogenous insulin within 2 years post-transplant. Therefore, there is an urgent requirement for the development of new strategies to prevent islet rejection. In this study, we explored the importance of human regulatory T cells in the control of islets allograft rejection. We developed a pre-clinical model of human islet transplantation by reconstituting NOD-scid IL2rγnull mice with cord blood-derived human CD34+ stem cells and demonstrated that although the engrafted human immune system mediated the rejection of human islets, their survival was significantly prolonged following adoptive transfer of ex vivo expanded human Tregs. Mechanistically, Tregs inhibited the infiltration of innate immune cells and CD4+ T cells into the graft by down-regulating the islet graft-derived monocyte chemoattractant protein-1. Our findings might contribute to the development of clinical strategies for Treg therapy to control human islet rejection. We also show for the first time that CD34+ cells-reconstituted NOD-scid IL2rγnull mouse model could be beneficial for investigating human innate immunity in vivo.

  13. Profound defects in pupillary responses to light in TRPM-channel null mice: a role for TRPM channels in non-image-forming photoreception.

    PubMed

    Hughes, Steven; Pothecary, Carina A; Jagannath, Aarti; Foster, Russell G; Hankins, Mark W; Peirson, Stuart N

    2012-01-01

    TRPM1 is a spontaneously active non-selective cation channel that has recently been shown to play an important role in the depolarizing light responses of ON bipolar cells. Consistent with this role, mutations in the TRPM1 gene have been identified as a principal cause of congenital stationary night blindness. However, previous microarray studies have shown that Trpm1 and Trpm3 are acutely regulated by light in the eyes of mice lacking rods and cones (rd/rd cl), a finding consistent with a role in non-image-forming photoreception. In this study we show that pupillary light responses are significantly attenuated in both Trpm1(-/-) and Trpm3(-/-) animals. Trpm1(-/-) mice exhibit a profound deficit in the pupillary response that is far in excess of that observed in mice lacking rods and cones (rd/rd cl) or melanopsin, and cannot be explained by defects in bipolar cell function alone. Immunolocalization studies suggest that TRPM1 is expressed in ON bipolar cells and also a subset of cells in the ganglion cell layer, including melanopsin-expressing photosensitive retinal ganglion cells (pRGCs). We conclude that, in addition to its role in bipolar cell signalling, TRPM1 is involved in non-image-forming responses to light and may perform a functional role within pRGCs. By contrast, TRPM3(-/-) mice display a more subtle pupillary phenotype with attenuated responses under bright light and dim light conditions. Expression of TRPM3 is detected in Muller cells and the ciliary body but is absent from pRGCs, and thus our data support an indirect role for TRPM3 in pupillary light responses.

  14. Comparison of effects of p53 null and gain-of-function mutations on salivary tumors in MMTV-Hras transgenic mice.

    PubMed

    Jiang, Dadi; Dumur, Catherine I; Massey, H Davis; Ramakrishnan, Viswanathan; Subler, Mark A; Windle, Jolene J

    2015-01-01

    p53 is an important tumor suppressor gene which is mutated in ~50% of all human cancers. Some of these mutants appear to have acquired novel functions beyond merely losing wild-type functions. To investigate these gain-of-function effects in vivo, we generated mice of three different genotypes: MMTV-Hras/p53(+/+), MMTV-Hras/p53(-/-), and MMTV-Hras/p53R172H/R172H. Salivary tumors from these mice were characterized with regard to age of tumor onset, tumor growth rates, cell cycle distribution, apoptotic levels, tumor histopathology, as well as response to doxorubicin treatment. Microarray analysis was also performed to profile gene expression. The MMTV-Hras/p53(-/-) and MMTV-Hras/p53R172H/R172H mice displayed similar properties with regard to age of tumor onset, tumor growth rates, tumor histopathology, and response to doxorubicin, while both groups were clearly distinct from the MMTV-Hras/p53(+/+) mice by these measurements. In addition, the gene expression profiles of the MMTV-Hras/p53(-/-) and MMTV-Hras/p53(R172H/R172H) tumors were tightly clustered, and clearly distinct from the profiles of the MMTV-Hras/p53(+/+) tumors. Only a small group of genes showing differential expression between the MMTV-Hras/p53(-/-) and MMTV-Hras/p53(R172H/R172H) tumors, that did not appear to be regulated by wild-type p53, were identified. Taken together, these results indicate that in this MMTV-Hras-driven salivary tumor model, the major effect of the p53 R172H mutant is due to the loss of wild-type p53 function, with little or no gain-of-function effect on tumorigenesis, which may be explained by the tissue- and tumor type-specific properties of this gain-of-function mutant of p53.

  15. Comparison of Effects of p53 Null and Gain-of-Function Mutations on Salivary Tumors in MMTV-Hras Transgenic Mice

    PubMed Central

    Jiang, Dadi; Dumur, Catherine I.; Massey, H. Davis; Ramakrishnan, Viswanathan; Subler, Mark A.; Windle, Jolene J.

    2015-01-01

    p53 is an important tumor suppressor gene which is mutated in ~50% of all human cancers. Some of these mutants appear to have acquired novel functions beyond merely losing wild-type functions. To investigate these gain-of-function effects in vivo, we generated mice of three different genotypes: MMTV-Hras/p53+/+, MMTV-Hras/p53-/-, and MMTV-Hras/p53R172H/R172H. Salivary tumors from these mice were characterized with regard to age of tumor onset, tumor growth rates, cell cycle distribution, apoptotic levels, tumor histopathology, as well as response to doxorubicin treatment. Microarray analysis was also performed to profile gene expression. The MMTV-Hras/p53-/- and MMTV-Hras/p53R172H/R172H mice displayed similar properties with regard to age of tumor onset, tumor growth rates, tumor histopathology, and response to doxorubicin, while both groups were clearly distinct from the MMTV-Hras/p53+/+ mice by these measurements. In addition, the gene expression profiles of the MMTV-Hras/p53-/- and MMTV-Hras/p53R172H/R172H tumors were tightly clustered, and clearly distinct from the profiles of the MMTV-Hras/p53+/+ tumors. Only a small group of genes showing differential expression between the MMTV-Hras/p53-/- and MMTV-Hras/p53R172H/R172H tumors, that did not appear to be regulated by wild-type p53, were identified. Taken together, these results indicate that in this MMTV-Hras-driven salivary tumor model, the major effect of the p53 R172H mutant is due to the loss of wild-type p53 function, with little or no gain-of-function effect on tumorigenesis, which may be explained by the tissue- and tumor type-specific properties of this gain-of-function mutant of p53. PMID:25695772

  16. Neuronal Activity-Induced Sterol Regulatory Element Binding Protein-1 (SREBP1) is Disrupted in Dysbindin-Null Mice-Potential Link to Cognitive Impairment in Schizophrenia.

    PubMed

    Chen, Yong; Bang, Sookhee; McMullen, Mary F; Kazi, Hala; Talbot, Konrad; Ho, Mei-Xuan; Carlson, Greg; Arnold, Steven E; Ong, Wei-Yi; Kim, Sangwon F

    2017-04-01

    Schizophrenia is a chronic debilitating neuropsychiatric disorder that affects about 1 % of the population. Dystrobrevin-binding protein 1 (DTNBP1 or dysbindin) is one of the Research Domain Constructs (RDoC) associated with cognition and is significantly reduced in the brain of schizophrenia patients. To further understand the molecular underpinnings of pathogenesis of schizophrenia, we have performed microarray analyses of the hippocampi from dysbindin knockout mice, and found that genes involved in the lipogenic pathway are suppressed. Moreover, we discovered that maturation of a master transcriptional regulator for lipid synthesis, sterol regulatory element binding protein-1 (SREBP1) is induced by neuronal activity, and is required for induction of the immediate early gene ARC (activity-regulated cytoskeleton-associated protein), necessary for synaptic plasticity and memory. We found that nuclear SREBP1 is dramatically reduced in dysbindin-1 knockout mice and postmortem brain tissues from human patients with schizophrenia. Furthermore, activity-dependent maturation of SREBP1 as well as ARC expression were attenuated in dysbindin-1 knockout mice, and these deficits were restored by an atypical antipsychotic drug, clozapine. Together, results indicate an important role of dysbindin-1 in neuronal activity induced SREBP1 and ARC, which could be related to cognitive deficits in schizophrenia.

  17. Anaplerotic Triheptanoin Diet Enhances Mitochondrial Substrate Use to Remodel the Metabolome and Improve Lifespan, Motor Function, and Sociability in MeCP2-Null Mice

    PubMed Central

    Li, Qun; Degano, Alicia L.; Penati, Judith; Zhuo, Justin; Roe, Charles R.; Ronnett, Gabriele V.

    2014-01-01

    Rett syndrome (RTT) is an autism spectrum disorder (ASD) caused by mutations in the X-linked MECP2 gene that encodes methyl-CpG binding protein 2 (MeCP2). Symptoms range in severity and include psychomotor disabilities, seizures, ataxia, and intellectual disability. Symptom onset is between 6-18 months of age, a critical period of brain development that is highly energy-dependent. Notably, patients with RTT have evidence of mitochondrial dysfunction, as well as abnormal levels of the adipokines leptin and adiponectin, suggesting overall metabolic imbalance. We hypothesized that one contributor to RTT symptoms is energy deficiency due to defective nutrient substrate utilization by the TCA cycle. This energy deficit would lead to a metabolic imbalance, but would be treatable by providing anaplerotic substrates to the TCA cycle to enhance energy production. We show that dietary therapy with triheptanoin significantly increased longevity and improved motor function and social interaction in male mice hemizygous for Mecp2 knockout. Anaplerotic therapy in Mecp2 knockout mice also improved indicators of impaired substrate utilization, decreased adiposity, increased glucose tolerance and insulin sensitivity, decreased serum leptin and insulin, and improved mitochondrial morphology in skeletal muscle. Untargeted metabolomics of liver and skeletal muscle revealed increases in levels of TCA cycle intermediates with triheptanoin diet, as well as normalizations of glucose and fatty acid biochemical pathways consistent with the improved metabolic phenotype in Mecp2 knockout mice on triheptanoin. These results suggest that an approach using dietary supplementation with anaplerotic substrate is effective in improving symptoms and metabolic health in RTT. PMID:25299635

  18. Muscle-specific AMPK β1β2-null mice display a myopathy due to loss of capillary density in nonpostural muscles.

    PubMed

    Thomas, Melissa M; Wang, David C; D'Souza, Donna M; Krause, Matthew P; Layne, Andrew S; Criswell, David S; O'Neill, Hayley M; Connor, Michael K; Anderson, Judy E; Kemp, Bruce E; Steinberg, Gregory R; Hawke, Thomas J

    2014-05-01

    AMP-activated protein kinase (AMPK) is a master regulator of metabolism. While muscle-specific AMPK β1β2 double-knockout (β1β2M-KO) mice display alterations in metabolic and mitochondrial capacity, their severe exercise intolerance suggested a secondary contributor to the observed phenotype. We find that tibialis anterior (TA), but not soleus, muscles of sedentary β1β2M-KO mice display a significant myopathy (decreased myofiber areas, increased split and necrotic myofibers, and increased centrally nucleated myofibers. A mitochondrial- and fiber-type-specific etiology to the myopathy was ruled out. However, β1β2M-KO TA muscles displayed significant (P<0.05) increases in platelet aggregation and apoptosis within myofibers and surrounding interstitium (P<0.05). These changes correlated with a 45% decrease in capillary density (P<0.05). We hypothesized that the β1β2M-KO myopathy in resting muscle resulted from impaired AMPK-nNOSμ signaling, causing increased platelet aggregation, impaired vasodilation, and, ultimately, ischemic injury. Consistent with this hypothesis, AMPK-specific phosphorylation (Ser1446) of nNOSμ was decreased in β1β2M-KO compared to wild-type (WT) mice. The AMPK-nNOSμ relationship was further demonstrated by administration of 5-aminoimidazole-4-carboxamide 1-β-D-ribofuranoside (AICAR) to β1β2-MKO muscles and C2C12 myotubes. AICAR significantly increased nNOSμ phosphorylation and nitric oxide production (P<0.05) within minutes of administration in WT muscles and C2C12 myotubes but not in β1β2M-KO muscles. These findings highlight the importance of the AMPK-nNOSμ pathway in resting skeletal muscle.

  19. Plaque Size Is Decreased but M1 Macrophage Polarization and Rupture Related Metalloproteinase Expression Are Maintained after Deleting T-Bet in ApoE Null Mice

    PubMed Central

    Tsaousi, Aikaterini; Hayes, Elaine M.; Di Gregoli, Karina; Bond, Andrew R.; Bevan, Laura

    2016-01-01

    Background Thelper1 (Th1) lymphocytes have been previously implicated in atherosclerotic plaque growth but their role in plaque vulnerability to rupture is less clear. We investigated whether T-bet knockout that prevents Th1 lymphocyte differentiation modulates classical (M1) macrophage activation or production of matrix degrading metalloproteinases (MMPs) and their tissue inhibitors, TIMPs. Methods & Results We studied the effect of T-bet deletion in apolipoproteinE (ApoE) knockout mice fed a high fat diet (HFD) or normal chow diet (ND). Transcript levels of M1/M2 macrophage polarization markers, selected MMPs and TIMPs were measured by RT-qPCR in macrophages isolated from subcutaneous granulomas or in whole aortae. Immunohistochemistry of aortic sinus (AS) and brachiocephalic artery (BCA) plaques was conducted to quantify protein expression of the same factors. Deletion of T-bet decreased mRNA for the M1 marker NOS-2 in granuloma macrophages but levels of M2 markers (CD206, arginase-1 and Ym-1), MMPs-2, -9, -12, -13, -14 and -19 or TIMPs-1 to -3 were unchanged. No mRNA differences were observed in aortic extracts from mice fed a HFD for 12 weeks. Moreover, AS and BCA plaques were similarly sized between genotypes, and had similar areas stained for NOS-2, COX-2, MMP-12 and MMP-14 proteins. T-bet deletion increased MMP-13, MMP-14 and arginase-1 in AS plaques. After 35 weeks of ND, T-bet deletion reduced the size of AS and BCA plaques but there were no differences in the percentage areas stained for M1 or M2 markers, MMPs-12, -13, -14, or TIMP-3. Conclusions Absence of Th1 lymphocytes is associated with reduced plaque size in ApoE knockout mice fed a normal but not high fat diet. In either case, M1 macrophage polarization and expression of several MMPs related to plaque instability are either maintained or increased. PMID:26886778

  20. Anaplerotic triheptanoin diet enhances mitochondrial substrate use to remodel the metabolome and improve lifespan, motor function, and sociability in MeCP2-null mice.

    PubMed

    Park, Min Jung; Aja, Susan; Li, Qun; Degano, Alicia L; Penati, Judith; Zhuo, Justin; Roe, Charles R; Ronnett, Gabriele V

    2014-01-01

    Rett syndrome (RTT) is an autism spectrum disorder (ASD) caused by mutations in the X-linked MECP2 gene that encodes methyl-CpG binding protein 2 (MeCP2). Symptoms range in severity and include psychomotor disabilities, seizures, ataxia, and intellectual disability. Symptom onset is between 6-18 months of age, a critical period of brain development that is highly energy-dependent. Notably, patients with RTT have evidence of mitochondrial dysfunction, as well as abnormal levels of the adipokines leptin and adiponectin, suggesting overall metabolic imbalance. We hypothesized that one contributor to RTT symptoms is energy deficiency due to defective nutrient substrate utilization by the TCA cycle. This energy deficit would lead to a metabolic imbalance, but would be treatable by providing anaplerotic substrates to the TCA cycle to enhance energy production. We show that dietary therapy with triheptanoin significantly increased longevity and improved motor function and social interaction in male mice hemizygous for Mecp2 knockout. Anaplerotic therapy in Mecp2 knockout mice also improved indicators of impaired substrate utilization, decreased adiposity, increased glucose tolerance and insulin sensitivity, decreased serum leptin and insulin, and improved mitochondrial morphology in skeletal muscle. Untargeted metabolomics of liver and skeletal muscle revealed increases in levels of TCA cycle intermediates with triheptanoin diet, as well as normalizations of glucose and fatty acid biochemical pathways consistent with the improved metabolic phenotype in Mecp2 knockout mice on triheptanoin. These results suggest that an approach using dietary supplementation with anaplerotic substrate is effective in improving symptoms and metabolic health in RTT.

  1. Angiotensin(1-7) and ACE2, “The Hot Spots” of Renin-Angiotensin System, Detected in the Human Aqueous Humor

    PubMed Central

    Holappa, Mervi; Valjakka, Jarkko; Vaajanen, Anu

    2015-01-01

    Background: The main purpose of the study was to establish whether essential components of the renin-angiotensin system (RAS) exist in the human aqueous humor. Methods: Forty-five patients ≥ 60 (74±7) years of age undergoing cataract surgery at Tampere University Hospital were randomly selected for the prospective study. The exclusion criterion was the use of oral antihypertensive medicine acting via renin-angiotensin system. Aqueous humor samples were taken at the beginning of normal cataract extraction. The samples were frozen and stored at -80 °C. The concentrations of intraocular endogenous RAS components Ang(1-7), ACE2, and ACE1 were measured using ELISA. Results: Concentration medians of Ang(1-7), ACE2, and ACE1 in the aqueous humor were: Ang(1-7) 4.08 ng/ml, ACE2 2.32 ng/ml and ACE1 0.35 ng/ml. The concentrations were significantly higher in glaucomatous than in non-glaucomatous eyes, ACE1 (p=0.014) and Ang(1-7) (p=0.026) vs non-glaucomatous eyes. Conclusions: Ang(1-7), ACE2 and ACE1 are found in the human aqueous humor. The observations are consistent with the conception that local tissue-RAS exists in the human eye and it might have a role in the control of intraocular pressure. PMID:25926900

  2. Nivolumab and Urelumab Enhance Antitumor Activity of Human T Lymphocytes Engrafted in Rag2-/-IL2Rγnull Immunodeficient Mice.

    PubMed

    Sanmamed, Miguel F; Rodriguez, Inmaculada; Schalper, Kurt A; Oñate, Carmen; Azpilikueta, Arantza; Rodriguez-Ruiz, Maria E; Morales-Kastresana, Aizea; Labiano, Sara; Pérez-Gracia, Jose L; Martín-Algarra, Salvador; Alfaro, Carlos; Mazzolini, Guillermo; Sarno, Francesca; Hidalgo, Manuel; Korman, Alan J; Jure-Kunkel, Maria; Melero, Ignacio

    2015-09-01

    A current pressing need in cancer immunology is the development of preclinical model systems that are immunocompetent for the study of human tumors. Here, we report the development of a humanized murine model that can be used to analyze the pharmacodynamics and antitumor properties of immunostimulatory monoclonal antibodies (mAb) in settings where the receptors targeted by the mAbs are expressed. Human lymphocytes transferred into immunodeficient mice underwent activation and redistribution to murine organs, where they exhibited cell-surface expression of hCD137 and hPD-1. Systemic lymphocyte infiltrations resulted in a lethal CD4(+) T cell-mediated disease (xenograft-versus-host disease), which was aggravated when murine subjects were administered clinical-grade anti-hCD137 (urelumab) and anti-hPD-1 (nivolumab). In mice engrafted with human colorectal HT-29 carcinoma cells and allogeneic human peripheral blood mononuclear cells (PBMC), or with a patient-derived gastric carcinoma and PBMCs from the same patient, we found that coadministration of urelumab and nivolumab was sufficient to significantly slow tumor growth. Correlated with this result were increased numbers of activated human T lymphocytes producing IFNγ and decreased numbers of human regulatory T lymphocytes in the tumor xenografts, possibly explaining the efficacy of the therapeutic regimen. Our results offer a proof of concept for the use of humanized mouse models for surrogate efficacy and histology investigations of immune checkpoint drugs and their combinations.

  3. Brain propagation of transduced α-synuclein involves non-fibrillar protein species and is enhanced in α-synuclein null mice.

    PubMed

    Helwig, Michael; Klinkenberg, Michael; Rusconi, Raffaella; Musgrove, Ruth E; Majbour, Nour K; El-Agnaf, Omar M A; Ulusoy, Ayse; Di Monte, Donato A

    2016-03-01

    Aggregation and neuron-to-neuron transmission are attributes of α-synuclein relevant to its pathogenetic role in human synucleinopathies such as Parkinson's disease. Intraparenchymal injections of fibrillar α-synuclein trigger widespread propagation of amyloidogenic protein species via mechanisms that require expression of endogenous α-synuclein and, possibly, its structural corruption by misfolded conformers acting as pathological seeds. Here we describe another paradigm of long-distance brain diffusion of α-synuclein that involves inter-neuronal transfer of monomeric and/or oligomeric species and is independent of recruitment of the endogenous protein. Targeted expression of human α-synuclein was induced in the mouse medulla oblongata through an injection of viral vectors into the vagus nerve. Enhanced levels of intra-neuronal α-synuclein were sufficient to initiate its caudo-rostral diffusion that likely involved at least one synaptic transfer and progressively reached specific brain regions such as the locus coeruleus, dorsal raphae and amygdala in the pons, midbrain and forebrain. Transfer of human α-synuclein was compared in two separate lines of α-synuclein-deficient mice versus their respective wild-type controls and, interestingly, lack of endogenous α-synuclein expression did not counteract diffusion but actually resulted in a more pronounced and advanced propagation of exogenous α-synuclein. Self-interaction of adjacent molecules of human α-synuclein was detected in both wild-type and mutant mice. In the former, interaction of human α-synuclein with mouse α-synuclein was also observed and might have contributed to differences in protein transmission. In wild-type and α-synuclein-deficient mice, accumulation of human α-synuclein within recipient axons in the pons, midbrain and forebrain caused morphological evidence of neuritic pathology. Tissue sections from the medulla oblongata and pons were stained with different antibodies recognizing

  4. A Novel Arginase Inhibitor Derived from Scutellavia indica Restored Endothelial Function in ApoE-Null Mice Fed a High-Cholesterol Diet.

    PubMed

    Hwang, Hye Mi; Lee, Jeong Hyung; Min, Byung Sun; Jeon, Byeong Hwa; Hoe, Kwang Lae; Kim, Young Myeong; Ryoo, Sungwoo

    2015-10-01

    Elevated endothelial arginase activity decreases nitric oxide (NO) production by competing with the substrate l-arginine, previously reported, and reciprocally regulating endothelial nitric oxide synthase (eNOS) activity. Thus, arginase inhibitors may help treat vascular diseases associated with endothelial dysfunction. A screening of metabolites from medicinal plants revealed that (2S)-5,2',5'-trihydroxy-7,8-dimethoxy flavanone (TDF) was a noncompetitive inhibitor of arginase. We investigated whether TDF reciprocally regulated endothelial NO production and its possible mechanism. TDF noncompetitively inhibited arginase I and II activity in a dose-dependent manner. TDF incubation decreased arginase activity and increased NO production in human umbilical vein endothelial cells and isolated mouse aortic vessels and reduced reactive oxygen species (ROS) generation in the endothelium of the latter. These TDF-mediated effects were associated with increased eNOS phosphorylation and dimerization but not with changes in protein content. Endothelium-dependent vasorelaxant responses to acetylcholine (Ach) were significantly increased in TDF-incubated aortic rings and attenuated by incubation with soluble guanylyl cyclase inhibitor. Phenylephrine-induced vasoconstrictor responses were markedly attenuated in TDF-treated vessels from wild-type mice. In atherogenic-prone ApoE(-/-) mice, TDF attenuated the high-cholesterol diet (HCD)-induced increase in arginase activity, which was accompanied by restoration of NO production and reduction of ROS generation. TDF incubation induced eNOS dimerization and phosphorylation at Ser1177. In addition, TDF improved Ach-dependent vasorelaxation responses and attenuated U46619-dependent contractile responses but did not change sodium nitroprusside-induced vasorelaxation or N-NAME-induced vasoconstriction. The findings suggest that TDF may help treat cardiovascular diseases by reducing pathophysiology derived from HCD-mediated endothelial

  5. Nhe1 is a luminal Na+/H+ exchanger in mouse choroid plexus and is targeted to the basolateral membrane in Ncbe/Nbcn2-null mice.

    PubMed

    Damkier, Helle Hasager; Prasad, Vikram; Hübner, Christian Andreas; Praetorius, Jeppe

    2009-06-01

    The choroid plexus epithelium (CPE) secretes the major fraction of the cerebrospinal fluid (CSF). The Na(+)-HCO(3)(-) transporter Ncbe/Nbcn2 in the basolateral membrane of CPE cells is important for Na(+)-dependent pH(i) increases and probably for CSF secretion. In the current study, the anion transport inhibitor DIDS had no effect on the residual pH(i) recovery in acidified CPE from Ncbe/Nbcn2 knockout mouse by 2',7'-bis(2-carboxyethyl)-5(6)-carboxyfluorescein (BCECF)-fluorescence microscopy in the presence of CO(2)/HCO(3)(-) (Ncbe/Nbcn2-ko+DIDS 109% of control, P = 0.76, n = 5). Thus Ncbe/Nbcn2 mediates the DIDS-sensitive Na(+)-dependent pH(i) recovery in the CPE. The Na(+)/H(+) exchanger-1 Nhe1 is proposed to mediate similar functions as Ncbe/Nbcn2 in CPE. Here, we immunolocalize the Nhe1 protein to the luminal membrane domain in mouse and human CPE. The Na(+)-dependent pH(i) recovery of Nhe1 wild-type (Nhe1-wt) mice in the absence of CO(2)/HCO(3)(-) was abolished in the Nhe1 knockout CPE (Nhe1-ko 0.37% of Nhe1-wt, P = 0.0007, n = 5). In Ncbe/Nbcn2-ko mice, Nhe1 was targeted to the basolateral membrane. Nevertheless, the luminal Na(+)-dependent pH(i) recovery was increased in Ncbe/Nbcn2-ko compared with wild-type littermates (Nhe1-ko 146% of Nhe1-wt, P = 0.007, n = 5). Whereas the luminal Nhe activity was inhibited by the Nhe blocker EIPA (10 microM) in the Ncbe/Nbcn2-wt, it was insensitive to the inhibitor in Ncbe/Nbcn2-ko (Ncbe/Nbcn2-ko+EIPA 100% of control, P = 0.98, n = 5). This indicates that a luminal EIPA-insensitive Nhe was induced in Ncbe/Nbcn2-ko CPE and that EIPA-sensitive Nhe activity was basolateral. The Nhe1 translocation in Ncbe/Nbcn2-ko CPE may reflect a compensatory response, which provides the cells with better means of regulating pH(i) or transporting Na(+) after Ncbe/Nbcn2 disruption.

  6. Endothelial nitric oxide synthase activation through obacunone-dependent arginase inhibition restored impaired endothelial function in ApoE-null mice.

    PubMed

    Yoon, Jeongyeon; Park, Minjin; Lee, Jeong hyung; Min, Byung Sun; Ryoo, Sungwoo

    2014-03-01

    Endothelial arginase constrains the activity of endothelial nitric oxide synthase (eNOS) by substrate depletion and reduces nitric oxide bioavailability. During the screening course of arginase inhibitor, we found obacunone as an arginase inhibitor. We tested the hypothesis that obacunone regulates vascular endothelial NO production. Obacunone incubation inhibited arginase I and II activities in liver and kidney lysates, respectively, in dose-dependent manner. Obacunone reciprocally increased nitrite/nitrate (NOx) production in HUVECs. In isolated aortic rings, obacunone increased intracellular l-arginine concentration and enhanced eNOS coupling, leading to increased NO and decreased superoxide production, with no changes in protein expression. Vasoconstriction response to U46619 was attenuated in obacunone-treated aortic vessels compared to that in untreated vessels. Endothelium-dependent vasorelaxant response to acetylcholine was significantly increased in obacunone-treated vessels and was modulated by the NO-dependent signaling cascade. The dose-dependent vasorelaxant response to Ach was reduced in the aortic vessels of ApoE-/- mice fed a high-cholesterol diet. Obacunone incubation increased vasorelaxation to the level of a WT mouse, although the endothelium-independent response to sodium nitroprusside was identical among the groups. Therefore, obacunone may help treat cardiovascular diseases derived from endothelial dysfunction and may be useful for designing pharmaceutical compounds.

  7. Interferometric Remapped Array Nulling

    NASA Astrophysics Data System (ADS)

    Abe, L.; Aristidi, E.; Vakili, F.; Domiciano, A.

    We present an interferometric beam recombination technique which allows achromatic and direct true imaging of targets at very high angular resolution. This technique intrinsically overcomes the main problems of Labeyrie's hypertelescope design, and can be used in a nulling configuration. It is thus particularly well suited for high contrast imaging in the context of exo-planet search and characterization especially for future space-borne arrays. We present the concept on a formal basis, and discuss its instrumental implementation.

  8. Extremely high levels of estradiol and testosterone in a case of polycystic ovarian syndrome. Hormone and clinical similarities with the phenotype of the alpha estrogen receptor null mice.

    PubMed

    Bartolone, L; Smedile, G; Arcoraci, V; Trimarchi, F; Benvenga, S

    2000-01-01

    A 19-year-old nulliparous hirsute woman was evaluated for the very high serum levels of testosterone (T) and estradiol (E2) measured in an outside laboratory. Menarche had occurred at 11 years and was followed by regular menses. We confirmed the high levels of T (9-16 ng/ml, nv 0.2-0.8) and E2 (>1,000 pg/ml, nv 30-120). LH and FSH were consistently high (73-118 mU/l and 18-29 mU/l, respectively; LH/FSH ratio=4.1-4.7) and responsive to iv GnRH (LH baseline=118 mU/I, 30 min=290; FSH baseline=25 mU/l, 30 min=46). The unstimulated values contrasted with those (LH=12, FSH=8 mU/I) measured in the outside laboratory, suggesting antigenically anomalous gonadotropins. 17-OH-progesterone was normal (0.5 ng/ml). After 1 mg dexamethasone, serum cortisol was normally suppressed (24-->0.4 microg/dl), T declined minimally (9-->8.6 ng/ml) and E2 remained high (>1,000 pg/ml). An exploratory laparotomy was performed, and two enlarged ovaries with multiple cysts as in a typical polycystic ovarian syndrome (PCOS) were seen. Before the wedge resection of the ovaries, hormones were assayed in the ovary veins (right ovary: T=30 ng/ml, Pg=17 ng/ml, E2=>5,000 pg/ml; left: T=14 ng/ml, Pg=14 ng/ml, E2=>5,000 pg/ml). Histologically, the follicle cysts showed luteinization of the theca interna; there was no evidence for ovary tumor in either ovary. After 21 days of 35 microg ethynyl-E2+2 mg cyproterone acetate (CA), E2=3,000 pg/ml, T=1.4 ng/ml, LH=10.5 mU/l and FSH=4.1 mU/I. After three cycles of the said therapy (but with 50 mg CA in the first 10 days of each cycle), E2 was 1,600 pg/ml, T 1.7 ng/ml, LH 7.1 and FSH 4.6 mU/I. Based on similarities with the phenotype of the alpha estrogen receptor knockout female mice (alphaERKO), one possible explanation for the puzzling clinical and biochemical picture of our patient is resistance of (alphaER to estrogens. This is the first case of PCOS with extremely high E2 and T. Thus, the differential diagnosis of high levels of E2 +/- T should include

  9. Chronic AT2 receptor activation increases renal ACE2 activity, attenuates AT1 receptor function and blood pressure in obese Zucker rats.

    PubMed

    Ali, Quaisar; Wu, Yonnie; Hussain, Tahir

    2013-11-01

    Abnormal regulation of the renin angiotensin system such as enhanced renal AT1R function and reduced ACE2 activity contributes to obesity-related hypertension. Here, we tested whether long-term AT2R activation affects renal function in obesity using lean and obese Zucker rats treated with the AT2R agonist CGP42112A for 2 weeks. This caused blood pressure to decrease by 13 mm Hg, which was associated with increased urinary sodium excretion in the obese rats. Cortical ACE2 expression and activity, the Mas receptor (MasR), and its ligand angiotensin-(1-7) were all increased in CGP-treated obese compared with control rats. Candesartan-induced natriuresis, a measure of AT₁R function, was reduced but cortical AT₁R expression and angiotensin II levels were similar in CGP-treated obese compared with control rats. Renin and AT2R expression in obese rats was not affected by CGP treatment. In HK-2 cells in vitro, CGP treatment caused increased ACE2 activity and MasR levels but decreased AT₁R levels and renin activity. Thus, long-term AT2R activation shifts the opposing arms of renin angiotensin system and contributes to natriuresis and blood pressure reduction in obese animals. Our study highlights the importance of AT2R as a target for treating obesity-related hypertension.

  10. Radiative Flux Changes by Aerosols from North America, Europe, and Africa over the Atlantic Ocean: Measurements and Calculations from TARFOX and ACE-2

    NASA Technical Reports Server (NTRS)

    Russell, P. B.; Hignett, P.; Livingston, J. M.; Schmid, B.; Chien, A.; Bergstrom, R.; Durkee, P. A.; Hobbs, P. V.; Bates, T. S.; Quinn, P. K.; Condon, Estelle (Technical Monitor)

    1998-01-01

    Aerosol effects on atmospheric radiative fluxes provide a forcing function that is a major source of uncertainty in understanding the past climate and predicting climate change. To help reduce this uncertainty, the 1996 Tropospheric Aerosol Radiative Forcing Experiment (TARFOX) and the 1997 second Aerosol Characterization Experiment (ACE-2) measured the properties and radiative effects of American, European, and African aerosols over the Atlantic. In TARFOX, radiative fluxes and microphysics of the American aerosol were measured from the UK C-130 while optical depth spectra, aerosol composition, and other properties were measured by the University of Washington C-131A and the CIRPAS Pelican. Closure studies show that the measured flux changes agree with those derived from the aerosol measurements using several modelling approaches. The best-fit midvisible single-scatter albedos (approx. 0.89 to 0.93) obtained from the TARFOX flux comparisons are in accord with values derived by independent techniques. In ACE-2 we measured optical depth and extinction spectra for both European urban-marine aerosols and free-tropospheric African dust aerosols, using sunphotometers on the R/V Vodyanitskiy and the Pelican. Preliminary values for the radiative flux sensitivities (Delta Flux / Delta Optical depth) computed for ACE-2 aerosols (boundary layer and African dust) over ocean are similar to those found in TARFOX. Combining a satellite-derived optical depth climatology with the aerosol optical model validated for flux sensitivities in TARFOX provides first-cut estimates of aerosol-induced flux changes over the Atlantic Ocean.

  11. Rac-null leukocytes are associated with increased inflammation-mediated alveolar bone loss.

    PubMed

    Sima, Corneliu; Gastfreund, Shoshi; Sun, Chunxiang; Glogauer, Michael

    2014-02-01

    Periodontitis is characterized by altered host-biofilm interactions that result in irreversible inflammation-mediated alveolar bone loss. Genetic and epigenetic factors that predispose to ineffective control of biofilm composition and maintenance of tissue homeostasis are not fully understood. We elucidated how leukocytes affect the course of periodontitis in Rac-null mice. Mouse models of acute gingivitis and periodontitis were used to assess the early inflammatory response and patterns of chronicity leading to loss of alveolar bone due to inflammation in Rac-null mice. Leukocyte margination was differentially impaired in these mice during attachment in conditional Rac1-null (granulocyte/monocyte lineage) mice and during rolling and attachment in Rac2-null (all blood cells) mice. Inflammatory responses to subgingival ligatures, assessed by changes in peripheral blood differential leukocyte numbers, were altered in Rac-null compared with wild-type mice. In response to persistent subgingival ligature-mediated challenge, Rac-null mice had increased loss of alveolar bone with patterns of resorption characteristic of aggressive forms of periodontitis. These findings were partially explained by higher osteoclastic coverage of the bone-periodontal ligament interface in Rac-null compared with wild-type mice. In conclusion, this study demonstrates that leukocyte defects, such as decreased endothelial margination and tissue recruitment, are rate-limiting steps in the periodontal inflammatory process that lead to more aggressive forms of periodontitis.

  12. Interferometric Remapped Array Nulling

    NASA Astrophysics Data System (ADS)

    Vakili, F.; Aristidi, E.; Abe, L.; Lopez, B.

    2004-07-01

    This paper describes a method of beam-combination in the so-called hypertelescope imaging technique recently introduced by Labeyrie in optical interferometry. The method we propose is an alternative to the Michelson pupil reconfiguration that suffers from the loss of the classical object-image convolution relation. From elementary theory of Fourier optics we demonstrate that this problem can be solved by reconfiguring images instead of pupils. Imaging is performed in a combined pupil-plane where the point-source intensity distribution (PSID by comparison to the more commonly quoted point-spread function, PSF) tends towards a pseudo Airy disc for a sufficiently large number of telescopes. Our method is applicable to snap-shot imaging of extended sources with a field limited to the Airy pattern of single telescopes operated in a co-phased multi-aperture interferometric array. It thus allows to apply conveniently pupil plane coronagraphy. Our technique called Interferometric Remapped Array Nulling (IRAN) is particularly suitable for high dynamic imaging of extra-solar planetary companions or extra-galactic objects where long baseline interferometry would closely probe the central regions of AGNs for instance. We also discuss the application of IRAN to improve the performances of imaging and/or nulling interferometers like the full-fledged VLTI array or the DARWIN space-borne mission.

  13. High Na intake increases renal angiotensin II levels and reduces expression of the ACE2-AT2R-MasR axis in obese Zucker rats

    PubMed Central

    Samuel, Preethi; Ali, Quaisar; Sabuhi, Rifat; Wu, Yonnie

    2012-01-01

    High sodium intake is known to regulate the renal renin-angiotensin system (RAS) and is a risk factor for the pathogenesis of obesity-related hypertension. The complex nature of the RAS reveals that its various components may have opposing effects on natriuresis and blood pressure regulation. We hypothesized that high sodium intake differentially regulates and shifts a balance between opposing components of the renal RAS, namely, angiotensin-converting enzyme (ACE)-ANG II-type 1 ANG II receptor (AT1R) vs. AT2-ACE2-angiotensinogen (Ang) (1–7)-Mas receptor (MasR), in obesity. In the present study, we evaluated protein and/or mRNA expression of angiotensinogen, renin, AT1A/BR, ACE, AT2R, ACE2, and MasR in the kidney cortex following 2 wk of a 8% high-sodium (HS) diet in lean and obese Zucker rats. The expression data showed that the relative expression pattern of ACE and AT1BR increased, renin decreased, and ACE2, AT2R, and MasR remained unaltered in HS-fed lean rats. On the other hand, HS intake in obese rats caused an increase in the cortical expression of ACE, a decrease in ACE2, AT2R, and MasR, and no changes in renin and AT1R. The cortical levels of ANG II increased by threefold in obese rats on HS compared with obese rats on normal salt (NS), which was not different than in lean rats. The HS intake elevated mean arterial pressure in obese rats (27 mmHg) more than in lean rats (16 mmHg). This study suggests that HS intake causes a pronounced increase in ANG II levels and a reduction in the expression of the ACE2-AT2R-MasR axis in the kidney cortex of obese rats. We conclude that such changes may lead to the potentially unopposed function of AT1R, with its various cellular and physiological roles, including the contribution to the pathogenesis of obesity-related hypertension. PMID:22592638

  14. High Na intake increases renal angiotensin II levels and reduces expression of the ACE2-AT(2)R-MasR axis in obese Zucker rats.

    PubMed

    Samuel, Preethi; Ali, Quaisar; Sabuhi, Rifat; Wu, Yonnie; Hussain, Tahir

    2012-08-01

    High sodium intake is known to regulate the renal renin-angiotensin system (RAS) and is a risk factor for the pathogenesis of obesity-related hypertension. The complex nature of the RAS reveals that its various components may have opposing effects on natriuresis and blood pressure regulation. We hypothesized that high sodium intake differentially regulates and shifts a balance between opposing components of the renal RAS, namely, angiotensin-converting enzyme (ACE)-ANG II-type 1 ANG II receptor (AT(1)R) vs. AT(2)-ACE2-angiotensinogen (Ang) (1-7)-Mas receptor (MasR), in obesity. In the present study, we evaluated protein and/or mRNA expression of angiotensinogen, renin, AT(1A/B)R, ACE, AT(2)R, ACE2, and MasR in the kidney cortex following 2 wk of a 8% high-sodium (HS) diet in lean and obese Zucker rats. The expression data showed that the relative expression pattern of ACE and AT(1B)R increased, renin decreased, and ACE2, AT(2)R, and MasR remained unaltered in HS-fed lean rats. On the other hand, HS intake in obese rats caused an increase in the cortical expression of ACE, a decrease in ACE2, AT(2)R, and MasR, and no changes in renin and AT(1)R. The cortical levels of ANG II increased by threefold in obese rats on HS compared with obese rats on normal salt (NS), which was not different than in lean rats. The HS intake elevated mean arterial pressure in obese rats (27 mmHg) more than in lean rats (16 mmHg). This study suggests that HS intake causes a pronounced increase in ANG II levels and a reduction in the expression of the ACE2-AT(2)R-MasR axis in the kidney cortex of obese rats. We conclude that such changes may lead to the potentially unopposed function of AT(1)R, with its various cellular and physiological roles, including the contribution to the pathogenesis of obesity-related hypertension.

  15. Influence of Humidity on the Aerosol Scattering Coefficient and Its Effect on the Upwelling Radiance During ACE-2

    NASA Technical Reports Server (NTRS)

    Gasso, B. S.; Hegg, D. A.; Covert, D. S.; Collins, D.; Noone, K.; Oestroem, E.; Schmid, B.; Russell, P. B.; Livingston, J. M.; Durkee, P. A.; Jonsson, H.

    2000-01-01

    Aerosol scattering coefficients (sigma(sub sp)) have been measured over the ocean at different relative humidities (RH) as a function of attitude in the region surrounding the Canary Islands during the Second Aerosol Characterization Experiment (ACE-2) in June and July 1997. The data were collected by the University of Washington passive humidigraph (UWPH) mounted on the Pelican research aircraft. Concurrently, particle size distributions absorption coefficients and aerosol optical depth were measured throughout 17 flights. A parameterization of sigma(sub sp) as a function of RH was utilized to assess the impact of aerosol hydration on the upwelling radiance (normalized to the solar constant and cosine of zenith angle). The top of the atmosphere radiance signal was simulated at wavelengths corresponding to visible and near-infrared bands or the EOS-AM ("Terra") detectors, MODIS and MISR. The UWPH measured (sigma(sub sp)) at 2 RHs, one below and the other above ambient conditions. Ambient (sigma(sub sp)) was obtained by interpolation of these 2 measurements. The data were stratified in terms of 3 types of aerosols: Saharan dust, clean marine (marine boundary layer background) and polluted marine aerosols (i.e., 2- or 1-day old polluted aerosols advected from Europe). An empirical relation for the dependence of (sigma(sub sp)) on RH, defined by (sigma(sub sp))(RH) = k. ((1 - RH/100)(exp -gamma), was used with the hygroscopic exponent gamma derived from the data. The following gamma values were obtained for the 3 aerosol types: gamma(dust) = 0.23 +/- 0.05, gamma(clean marine) = 0.69 +/- 0.06 and gamma(polluted marine) = 0.57 + 0.06. Based on the measured (gamma)(s), the above equation was utilized to derive aerosol models with different hygroscopicities. The satellite simulation signal code 6S was used to compute the upwelling radiance corresponding to each of those aerosol models at several ambient humidities. For the pre-launch estimated precision of the sensors and

  16. Influence of Humidity On the Aerosol Scattering Coefficient and Its Effect on the Upwelling Radiance During ACE-2

    NASA Technical Reports Server (NTRS)

    Gasso, S.; Hegg, D. A.; Covert, D. S.; Collins, D.; Noone, K. J.; Oestroem, E.; Schmid, B.; Russell, P. B.; Livingston, J. M.; Durkee, P. A.

    2000-01-01

    Aerosol scattering coefficients (sigma(sub sp)) have been measured over the ocean at different relative humidities (RH) as a function of altitude in the region surrounding the Canary Islands during the Second Aerosol Characterization Experiment (ACE-2) in June and July 1997. The data were collected by the University of Washington passive humidigraph (UWPH) mounted on the Pelican research aircraft. Concurrently, particle size distributions, absorption coefficients and aerosol optical depth were measured throughout 17 flights. A parameterization of sigma(sub sp) as a function of RH was utilized to assess the impact of aerosol hydration on the upwelling radiance (normalized to the solar constant and cosine of zenith angle). The top of the atmosphere radiance signal was simulated at wavelengths corresponding to visible and near-infrared bands of the EOS (Earth Observing System) AM-1 (Terra) detectors, MODIS (Moderate Resolution Imaging Spectroradiometer) and MISR (Multi-angle Imaging Spectroradiometer). The UWPH measured sigma(sub sp) at two RHs, one below and the other above ambient conditions. Ambient sigma(sub sp) was obtained by interpolation of these two measurements. The data were stratified in terms of three types of aerosols: Saharan dust, clean marine (marine boundary layer background) and polluted marine aerosols (i.e., two- or one-day old polluted aerosols advected from Europe). An empirical relation for the dependence of sigma(sub sp) on RH, defined by sigma(sub sp)(RH) = k.(1 - RH/100)(sup gamma), was used with the hygroscopic exponent gamma derived from the data. The following gamma values were obtained for the 3 aerosol types: gamma(dust) = 0.23 +/- 0.05, gamma(clean marine) = 0.69 +/- 0.06 and gamma(polluted marine) = 0.57 +/- 0.06. Based on the measured gammas, the above equation was utilized to derive aerosol models with different hygroscopicities. The satellite simulation signal code 6S was used to compute the upwelling radiance corresponding to each

  17. Thermogenic characterization of ghrelin receptor null mice

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Ghrelin is the only known circulating orexigenic hormone that increases food intake and promotes adiposity, and these physiological functions of ghrelin are mediated through its receptor growth hormone secretagogue receptor (GHS-R). Ghrelin/GHS-R signaling plays a crucial role in energy homeostasis....

  18. Radiant Temperature Nulling Radiometer

    NASA Technical Reports Server (NTRS)

    Ryan, Robert (Inventor)

    2003-01-01

    A self-calibrating nulling radiometer for non-contact temperature measurement of an object, such as a body of water, employs a black body source as a temperature reference, an optomechanical mechanism, e.g., a chopper, to switch back and forth between measuring the temperature of the black body source and that of a test source, and an infrared detection technique. The radiometer functions by measuring radiance of both the test and the reference black body sources; adjusting the temperature of the reference black body so that its radiance is equivalent to the test source; and, measuring the temperature of the reference black body at this point using a precision contact-type temperature sensor, to determine the radiative temperature of the test source. The radiation from both sources is detected by an infrared detector that converts the detected radiation to an electrical signal that is fed with a chopper reference signal to an error signal generator, such as a synchronous detector, that creates a precision rectified signal that is approximately proportional to the difference between the temperature of the reference black body and that of the test infrared source. This error signal is then used in a feedback loop to adjust the reference black body temperature until it equals that of the test source, at which point the error signal is nulled to zero. The chopper mechanism operates at one or more Hertz allowing minimization of l/f noise. It also provides pure chopping between the black body and the test source and allows continuous measurements.

  19. The replication of a mouse adapted SARS-CoV in a mouse cell line stably expressing the murine SARS-CoV receptor mACE2 efficiently induces the expression of proinflammatory cytokines.

    PubMed

    Regla-Nava, Jose A; Jimenez-Guardeño, Jose M; Nieto-Torres, Jose L; Gallagher, Thomas M; Enjuanes, Luis; DeDiego, Marta L

    2013-11-01

    Infection of conventional mice with a mouse adapted (MA15) severe acute respiratory syndrome (SARS) coronavirus (CoV) reproduces many aspects of human SARS such as pathological changes in lung, viremia, neutrophilia, and lethality. However, established mouse cell lines highly susceptible to mouse-adapted SARS-CoV infection are not available. In this work, efficiently transfectable mouse cell lines stably expressing the murine SARS-CoV receptor angiotensin converting enzyme 2 (ACE2) have been generated. These cells yielded high SARS-CoV-MA15 titers and also served as excellent tools for plaque assays. In addition, in these cell lines, SARS-CoV-MA15 induced the expression of proinflammatory cytokines and IFN-β, mimicking what has been observed in experimental animal models infected with SARS-CoV and SARS patients. These cell lines are valuable tools to perform in vitro studies in a mouse cell system that reflects the species used for in vivo studies of SARS-CoV-MA15 pathogenesis.

  20. Null testing convex optical surfaces.

    PubMed

    Szulc, A

    1997-09-01

    A new test for convex optical surfaces is presented. It makes use of an auxiliary ellipsoidal mirror that is of approximately the same diameter as the convex mirror tested. The test is a null test of excellent precision. The auxiliary ellipsoid used is also tested in a null fashion, permitting good precision to be obtained.

  1. On the randomness of pulsar nulls

    NASA Astrophysics Data System (ADS)

    Redman, Stephen L.; Rankin, Joanna M.

    2009-05-01

    Pulsar nulling is not always a random process; most pulsars, in fact, null non-randomly. The Wald-Wolfowitz statistical runs test is a simple diagnostic that pulsar astronomers can use to identify pulsars that have non-random nulls. It is not clear at this point how the dichotomy in pulsar nulling randomness is related to the underlying nulling phenomenon, but its nature suggests that there are at least two distinct reasons that pulsars null.

  2. Differential regulation of the cellulase transcription factors XYR1, ACE2, and ACE1 in Trichoderma reesei strains producing high and low levels of cellulase.

    PubMed

    Portnoy, Thomas; Margeot, Antoine; Seidl-Seiboth, Verena; Le Crom, Stéphane; Ben Chaabane, Fadhel; Linke, Rita; Seiboth, Bernhard; Kubicek, Christian P

    2011-02-01

    Due to its capacity to produce large amounts of cellulases, Trichoderma reesei is increasingly being investigated for second-generation biofuel production from lignocellulosic biomass. The induction mechanisms of T. reesei cellulases have been described recently, but the regulation of the genes involved in their transcription has not been studied thoroughly. Here we report the regulation of expression of the two activator genes xyr1 and ace2, and the corepressor gene ace1, during the induction of cellulase biosynthesis by the inducer lactose in T. reesei QM 9414, a strain producing low levels of cellulase (low producer). We show that all three genes are induced by lactose. xyr1 was also induced by d-galactose, but this induction was independent of d-galactose metabolism. Moreover, ace1 was carbon catabolite repressed, whereas full induction of xyr1 and ace2 in fact required CRE1. Significant differences in these regulatory patterns were observed in the high-producer strain RUT C30 and the hyperproducer strain T. reesei CL847. These observations suggest that a strongly elevated basal transcription level of xyr1 and reduced upregulation of ace1 by lactose may have been important for generating the hyperproducer strain and that thus, these genes are major control elements of cellulase production.

  3. [Pyr1]Apelin-13(1–12) Is a Biologically Active ACE2 Metabolite of the Endogenous Cardiovascular Peptide [Pyr1]Apelin-13

    PubMed Central

    Yang, Peiran; Kuc, Rhoda E.; Brame, Aimée L.; Dyson, Alex; Singer, Mervyn; Glen, Robert C.; Cheriyan, Joseph; Wilkinson, Ian B.; Davenport, Anthony P.; Maguire, Janet J.

    2017-01-01

    Aims: Apelin is a predicted substrate for ACE2, a novel therapeutic target. Our aim was to demonstrate the endogenous presence of the putative ACE2 product [Pyr1]apelin-13(1–12) in human cardiovascular tissues and to confirm it retains significant biological activity for the apelin receptor in vitro and in vivo. The minimum active apelin fragment was also investigated. Methods and Results: [Pyr1]apelin-13 incubated with recombinant human ACE2 resulted in de novo generation of [Pyr1]apelin-13(1–12) identified by mass spectrometry. Endogenous [Pyr1]apelin-13(1–12) was detected by immunostaining in human heart and lung localized to the endothelium. Expression was undetectable in lung from patients with pulmonary arterial hypertension. In human heart [Pyr1]apelin-13(1–12) (pKi = 8.04 ± 0.06) and apelin-13(F13A) (pKi = 8.07 ± 0.24) competed with [125I]apelin-13 binding with nanomolar affinity, 4-fold lower than for [Pyr1]apelin-13 (pKi = 8.83 ± 0.06) whereas apelin-17 exhibited highest affinity (pKi = 9.63 ± 0.17). The rank order of potency of peptides to inhibit forskolin-stimulated cAMP was apelin-17 (pD2 = 10.31 ± 0.28) > [Pyr1]apelin-13 (pD2 = 9.67 ± 0.04) ≥ apelin-13(F13A) (pD2 = 9.54 ± 0.05) > [Pyr1]apelin-13(1–12) (pD2 = 9.30 ± 0.06). The truncated peptide apelin-13(R10M) retained nanomolar potency (pD2 = 8.70 ± 0.04) but shorter fragments exhibited low micromolar potency. In a β-arrestin recruitment assay the rank order of potency was apelin-17 (pD2 = 10.26 ± 0.09) >> [Pyr1]apelin-13 (pD2 = 8.43 ± 0.08) > apelin-13(R10M) (pD2 = 8.26 ± 0.17) > apelin-13(F13A) (pD2 = 7.98 ± 0.04) ≥ [Pyr1]apelin-13(1–12) (pD2 = 7.84 ± 0.06) >> shorter fragments (pD2 < 6). [Pyr1]apelin-13(1–12) and apelin-13(F13A) contracted human saphenous vein with similar sub-nanomolar potencies and [Pyr1]apelin-13(1–12) was a potent inotrope in paced mouse right ventricle and human atria. [Pyr1]apelin-13(1–12) elicited a dose-dependent decrease in blood

  4. North Atlantic Aerosol Single Scattering Albedos: TARFOX and ACE-2 Results and Their Relation to Radiative Effects Derived from Satellite Optical Depths

    NASA Technical Reports Server (NTRS)

    Russell, P. B.; Bergstrom, R. W.; Schmid, B.; Livingston, J. M.; Redemann, J.; Quinn, P. K.; Carrico, C. M.; Rood, M. J.

    2000-01-01

    Bergstrom and Russell estimated direct solar radiative flux changes caused by atmospheric aerosols over the mid-latitude North Atlantic Ocean under cloud-free and cloudy conditions. They excluded African dust aerosols, primarily by restricting calculations to latitudes 25-60 N. As inputs they used midvisible aerosol optical depth (AOD) maps derived from AVHRR satellite measurements and aerosol intensive properties determined primarily in the 1996 IGAC Troposheric Aerosol Radiative Forcing Observational Experiment (TARFOX). Those aerosol intensive properties, which included optical depth wavelength dependence and spectra of single scattering albedo (SSA) and scattering asymmetry parameter, were also checked against initial properties from the 1997 North Atlantic Aerosol Characterization Experiment (ACE 2). Bergstrom and Russell investigated the sensitivity of their derived flux changes to assumed input parameters, including midvisible AOD, SSA, and scattering asymmetry parameter. Although the sensitivity of net flux change at the tropopause to SSA was moderate over the ocean (e.g., a SSA uncertainty of 0.07 produced a flux-change uncertainty of 21%), the sensitivity over common land surfaces can be much larger. Also, flux changes within and below the aerosol layer, which affect atmospheric stability, heating rates, and cloud formation and persistence, are quite sensitive to aerosol SSA. Therefore, this paper focuses on the question: "What have we learned from TARFOX and ACE 2 regarding aerosol single scattering albedo?" Three techniques were used in TARFOX to determine midvisible SSA. One of these derived SSA as a best-fit parameter in comparing radiative flux changes measured by airborne pyranometer to those computed from aerosol properties. Another technique combined airborne measurements of aerosol scattering and absorption by nephelometer and absorption photometer. A third technique obtained SSA from best-fit complex refractive indices derived by comparing

  5. Null lifts and projective dynamics

    SciTech Connect

    Cariglia, Marco

    2015-11-15

    We describe natural Hamiltonian systems using projective geometry. The null lift procedure endows the tangent bundle with a projective structure where the null Hamiltonian is identified with a projective conic and induces a Weyl geometry. Projective transformations generate a set of known and new dualities between Hamiltonian systems, as for example the phenomenon of coupling-constant metamorphosis. We conclude outlining how this construction can be extended to the quantum case for Eisenhart–Duval lifts.

  6. Gravitational action with null boundaries

    NASA Astrophysics Data System (ADS)

    Lehner, Luis; Myers, Robert C.; Poisson, Eric; Sorkin, Rafael D.

    2016-10-01

    We present a complete discussion of the boundary term in the action functional of general relativity when the boundary includes null segments in addition to the more usual timelike and spacelike segments. We confirm that ambiguities appear in the contribution from a null segment, because it depends on an arbitrary choice of parametrization for the generators. We also show that similar ambiguities appear in the contribution from a codimension-two surface at which a null segment is joined to another (spacelike, timelike, or null) segment. The parametrization ambiguity can be tamed by insisting that the null generators be affinely parametrized; this forces each null contribution to the boundary action to vanish, but leaves intact the fredom to rescale the affine parameter by a constant factor on each generator. Once a choice of parametrization is made, the ambiguity in the joint contributions can be eliminated by formulating well-motivated rules that ensure the additivity of the gravitational action. Enforcing these rules, we calculate the time rate of change of the action when it is evaluated for a so-called "Wheeler-DeWitt patch" of a black hole in asymptotically anti de Sitter space. We recover a number of results cited in the literature, obtained with a less complete analysis.

  7. Identification of residues in the receptor-binding domain (RBD) of the spike protein of human coronavirus NL63 that are critical for the RBD-ACE2 receptor interaction.

    PubMed

    Lin, Han-Xin; Feng, Yan; Wong, Gillian; Wang, Liping; Li, Bei; Zhao, Xuesen; Li, Yan; Smaill, Fiona; Zhang, Chengsheng

    2008-04-01

    Human coronavirus NL63 (NL63), a member of the group I coronaviruses, may cause acute respiratory diseases in young children and immunocompromised adults. Like severe acute respiratory syndrome coronavirus (SARS-CoV), NL63 also employs the human angiotensin-converting enzyme 2 (hACE2) receptor for cellular entry. To identify residues in the spike protein of NL63 that are important for hACE2 binding, this study first generated a series of S1-truncated variants, examined their associations with the hACE2 receptor and subsequently mapped a minimal receptor-binding domain (RBD) that consisted of 141 residues (aa 476-616) towards the C terminus of the S1 domain. The data also demonstrated that the NL63 RBD bound to hACE2 more efficiently than its full-length counterpart and had a binding efficiency comparable to the S1 or RBD of SARS-CoV. A further series of RBD variants was generated using site-directed mutagenesis and random mutant library screening assays, and identified 15 residues (C497, Y498, V499, C500, K501, R518, R530, V531, G534, G537, D538, S540, E582, W585 and T591) that appeared to be critical for the RBD-hACE2 association. These critical residues clustered in three separate regions (designated RI, RII and RIII) inside the RBD, which may represent three receptor-binding sites. These results may help to delineate the molecular interactions between the S protein of NL63 and the hACE2 receptor, and may also enhance our understanding of the pathogenesis of NL63 and SARS-CoV.

  8. NULL convention floating point multiplier.

    PubMed

    Albert, Anitha Juliette; Ramachandran, Seshasayanan

    2015-01-01

    Floating point multiplication is a critical part in high dynamic range and computational intensive digital signal processing applications which require high precision and low power. This paper presents the design of an IEEE 754 single precision floating point multiplier using asynchronous NULL convention logic paradigm. Rounding has not been implemented to suit high precision applications. The novelty of the research is that it is the first ever NULL convention logic multiplier, designed to perform floating point multiplication. The proposed multiplier offers substantial decrease in power consumption when compared with its synchronous version. Performance attributes of the NULL convention logic floating point multiplier, obtained from Xilinx simulation and Cadence, are compared with its equivalent synchronous implementation.

  9. NULL Convention Floating Point Multiplier

    PubMed Central

    Ramachandran, Seshasayanan

    2015-01-01

    Floating point multiplication is a critical part in high dynamic range and computational intensive digital signal processing applications which require high precision and low power. This paper presents the design of an IEEE 754 single precision floating point multiplier using asynchronous NULL convention logic paradigm. Rounding has not been implemented to suit high precision applications. The novelty of the research is that it is the first ever NULL convention logic multiplier, designed to perform floating point multiplication. The proposed multiplier offers substantial decrease in power consumption when compared with its synchronous version. Performance attributes of the NULL convention logic floating point multiplier, obtained from Xilinx simulation and Cadence, are compared with its equivalent synchronous implementation. PMID:25879069

  10. Achronal averaged null energy condition

    SciTech Connect

    Graham, Noah; Olum, Ken D.

    2007-09-15

    The averaged null energy condition (ANEC) requires that the integral over a complete null geodesic of the stress-energy tensor projected onto the geodesic tangent vector is never negative. This condition is sufficient to prove many important theorems in general relativity, but it is violated by quantum fields in curved spacetime. However there is a weaker condition, which is free of known violations, requiring only that there is no self-consistent spacetime in semiclassical gravity in which ANEC is violated on a complete, achronal null geodesic. We indicate why such a condition might be expected to hold and show that it is sufficient to rule out closed timelike curves and wormholes connecting different asymptotically flat regions.

  11. Shortwave Radiative Fluxes, Solar-Beam Transmissions, and Aerosol Properties: TARFOX and ACE-2 Find More Absorption from Flux Radiometry than from Other Measurements

    NASA Technical Reports Server (NTRS)

    Russell, Philip B.; Redemann, J.; Schmid, B.; Livingston, J. M.; Bergstrom, R. W.; Ramirez, S. A.; Hipskind, R. Stephen (Technical Monitor)

    2001-01-01

    The Tropospheric Aerosol Radiative Forcing Observational Experiment (TARFOX) and the Second Aerosol Characterization Experiment (ACE-2) made simultaneous measurements of shortwave radiative fluxes, solar-beam transmissions, and the aerosols affecting those fluxes and transmissions. Besides the measured fluxes and transmissions, other obtained properties include aerosol scattering and absorption measured in situ at the surface and aloft; aerosol single scattering albedo retrieved from skylight radiances; and aerosol complex refractive index derived by combining profiles of backscatter, extinction, and size distribution. These measurements of North Atlantic boundary layer aerosols impacted by anthropogenic pollution revealed the following characteristic results: (1) Better agreement among different types of remote measurements of aerosols (e.g., optical depth, extinction, and backscattering from sunphotometers, satellites, and lidars) than between remote and in situ measurements; 2) More extinction derived from transmission measurements than from in situ measurements; (3) Larger aerosol absorption inferred from flux radiometry than from other measurements. When the measured relationships between downwelling flux and optical depth (or beam transmission) are used to derive best-fit single scattering albedos for the polluted boundary layer aerosol, both TARFOX and ACE-2 yield midvisible values of 0.90 +/- 0.04. The other techniques give larger single scattering albedos (i.e. less absorption) for the polluted boundary layer, with a typical result of 0.95 +/- 0.04. Although the flux-based results have the virtue of describing the column aerosol unperturbed by sampling, they are subject to questions about representativeness and other uncertainties (e.g., unknown gas absorption). Current uncertainties in aerosol single scattering albedo are large in terms of climate effects. They also have an important influence on aerosol optical depths retrieved from satellite radiances

  12. Attenuation of myocardial fibrosis with curcumin is mediated by modulating expression of angiotensin II AT1/AT2 receptors and ACE2 in rats

    PubMed Central

    Pang, Xue-Fen; Zhang, Li-Hui; Bai, Feng; Wang, Ning-Ping; Garner, Ron E; McKallip, Robert J; Zhao, Zhi-Qing

    2015-01-01

    Curcumin is known to improve cardiac function by balancing degradation and synthesis of collagens after myocardial infarction. This study tested the hypothesis that inhibition of myocardial fibrosis by curcumin is associated with modulating expression of angiotensin II (Ang II) receptors and angiotensin-converting enzyme 2 (ACE2). Male Sprague Dawley rats were subjected to Ang II infusion (500 ng/kg/min) using osmotic minipumps for 2 and 4 weeks, respectively, and curcumin (150 mg/kg/day) was fed by gastric gavage during Ang II infusion. Compared to the animals with Ang II infusion, curcumin significantly decreased the mean arterial blood pressure during the course of the observation. The protein level of the Ang II type 1 (AT1) receptor was reduced, and the Ang II type 2 (AT2) receptor was up-regulated, evidenced by an increased ratio of the AT2 receptor over the AT1 receptor in the curcumin group (1.2±0.02%) vs in the Ang II group (0.7±0.03%, P<0.05). These changes were coincident with less locally expressed AT1 receptor and enhanced AT2 receptor in the intracardiac vessels and intermyocardium. Along with these modulations, curcumin significantly decreased the populations of macrophages and alpha smooth muscle actin-expressing myofibroblasts, which were accompanied by reduced expression of transforming growth factor beta 1 and phosphorylated-Smad2/3. Collagen I synthesis was inhibited, and tissue fibrosis was attenuated, as demonstrated by less extensive collagen-rich fibrosis. Furthermore, curcumin increased protein level of ACE2 and enhanced its expression in the intermyocardium relative to the Ang II group. These results suggest that curcumin could be considered as an add-on therapeutic agent in the treatment of fibrosis-derived heart failure patient who is intolerant of ACE inhibitor therapy. PMID:26648693

  13. The atherogenic Scarb1 null mouse model shows a high bone mass phenotype.

    PubMed

    Martineau, Corine; Martin-Falstrault, Louise; Brissette, Louise; Moreau, Robert

    2014-01-01

    Scavenger receptor class B, type I (SR-BI), the Scarb1 gene product, is a receptor associated with cholesteryl ester uptake from high-density lipoproteins (HDL), which drives cholesterol movement from peripheral tissues toward the liver for excretion, and, consequently, Scarb1 null mice are prone to atherosclerosis. Because studies have linked atherosclerosis incidence with osteoporosis, we characterized the bone metabolism in these mice. Bone morphometry was assessed through microcomputed tomography and histology. Marrow stromal cells (MSCs) were used to characterize influence of endogenous SR-BI in cell functions. Total and HDL-associated cholesterol in null mice were increased by 32-60%, correlating with its role in lipoprotein metabolism. Distal metaphyses from 2- and 4-mo-old null mice showed correspondingly 46 and 37% higher bone volume fraction associated with a higher number of trabeculae. Histomorphometric analyses in 2-mo-old null male mice revealed 1.42-fold greater osteoblast surface, 1.37-fold higher percent mineralizing surface, and 1.69-fold enhanced bone formation rate. In vitro assays for MSCs from null mice revealed 37% higher proliferation rate, 48% more alkaline phosphatase activity, 70% greater mineralization potential and a 2-fold osterix (Sp7) expression, yet a 0.5-fold decrease in caveolin-1 (Cav1) expression. Selective uptake levels of HDL-associated cholesteryl oleate and estradiol were similar between MSC from wild-type and Scarb1 null mice, suggesting that its contribution to this process is not its main role in these cells. However, Scarb1 knockout stunted the HDL-dependent regulation of Cav1 genic expression. Scarb1 null mice are not prone to osteoporosis but show higher bone mass associated with enhanced bone formation.

  14. Proliferation, differentiation and apoptosis in connexin43-null osteoblasts

    NASA Technical Reports Server (NTRS)

    Furlan, F.; Lecanda, F.; Screen, J.; Civitelli, R.

    2001-01-01

    Osteoblasts are highly coupled by gap junctions formed primarily by connexin43 (Cx43). We have shown that interference with Cx43 expression or function disrupts transcriptional regulation of osteoblast genes, and that deletion of Cx43 in the mouse causes skeletal malformations, delayed mineralization, and osteoblast dysfunction. Here, we studied the mechanisms by which genetic deficiency of Cx43 alters osteoblast development. While cell proliferation rates were similar in osteoblastic cells derived from calvaria of Cx43-null and wild type mice, camptothecin-induced apoptosis was 3-fold higher in mutant compared to wild type osteoblasts. When grown in mineralizing medium, Cx43-null cells were able to produce mineralized matrix but it took one week longer to reach the same mineralization levels as in normal cells. Likewise, expression of alkaline phosphatase activity per cell--a marker of osteoblast differentiation--was maximal only 2 weeks later in Cx43-null relative to wild-type cells. These observations suggest that Cx43 is important for a normal and timely development of the osteoblastic phenotype. Delayed differentiation and increase programmed cell death may explain the skeletal phenotype of Cx43-null mice.

  15. Effects of Local Heart Irradiation in a Glutathione S-Transferase Alpha 4-Null Mouse Model.

    PubMed

    Boerma, Marjan; Singh, Preeti; Sridharan, Vijayalakshmi; Tripathi, Preeti; Sharma, Sunil; Singh, Sharda P

    2015-06-01

    Glutathione S-transferase alpha 4 (GSTA4-4) is one of the enzymes responsible for the removal of 4-hydroxynonenal (4-HNE), an electrophilic product of lipid peroxidation in cellular membranes during oxidative stress. 4-HNE is a direct activator of nuclear factor (erythroid-derived 2)-like 2 (Nrf2), a transcription factor with many target genes encoding antioxidant and anti-electrophile enzymes. We have previously shown that Gsta4-null mice on a 129/Sv background exhibited increased activity of Nrf2 in the heart. Here we examined the sensitivity of this Gsta4-null mouse model towards cardiac function and structure loss due to local heart irradiation. Male Gsta4-null and wild-type mice were exposed to a single X-ray dose of 18 Gy to the heart. Six months after irradiation, immunohistochemical staining for respiratory complexes 2 and 5 indicated that radiation exposure had caused most pronounced alterations in mitochondrial morphology in Gsta4-null mice. On the other hand, wild-type mice showed a decline in cardiac function and an increase in plasma levels of troponin-I, while no such changes were observed in Gsta4-null mice. Radiation-induced Nrf2-target gene expression only in Gsta4-null mice. In conclusion, although loss of GSTA4-4 led to enhanced susceptibility of cardiac mitochondria to radiation-induced loss of morphology, cardiac function was preserved in Gsta4-null mice. We propose that this protection against cardiac function loss may occur, at least in part, by upregulation of the Nrf2 pathway.

  16. Fibrosis and inflammation are greater in muscles of beta-sarcoglycan-null mouse than mdx mouse.

    PubMed

    Gibertini, Sara; Zanotti, Simona; Savadori, Paolo; Curcio, Maurizio; Saredi, Simona; Salerno, Franco; Andreetta, Francesca; Bernasconi, Pia; Mantegazza, Renato; Mora, Marina

    2014-05-01

    The Sgcb-null mouse, with knocked-down β-sarcoglycan, develops severe muscular dystrophy as in type 2E human limb girdle muscular dystrophy. The mdx mouse, lacking dystrophin, is the most used model for Duchenne muscular dystrophy (DMD). Unlike DMD, the mdx mouse has mild clinical features and shows little fibrosis in limb muscles. To characterize ECM protein deposition and the progression of muscle fibrosis, we evaluated protein and transcript levels of collagens I, III and VI, decorin, and TGF-β1, in quadriceps and diaphragm, at 2, 4, 8, 12, 26, and 52 weeks in Sgcb-null mice, and protein levels at 12, 26, and 52 weeks in mdx mice. In Sgcb-null mice, severe morphological disruption was present from 4 weeks in both quadriceps and diaphragm, and included conspicuous deposition of extracellular matrix components. Histopathological features of Sgcb-null mouse muscles were similar to those of age-matched mdx muscles at all ages examined, but, in the Sgcb-null mouse, the extent of connective tissue deposition was generally greater than mdx. Furthermore, in the Sgcb-null mouse, the amount of all three collagen isoforms increased steadily, while, in the mdx, they remained stable. We also found that, at 12 weeks, macrophages were significantly more numerous in mildly inflamed areas of Sgcb-null quadriceps compared to mdx quadriceps (but not in highly inflamed regions), while, in the diaphragm, macrophages did not differ significantly between the two models, in either region. Osteopontin mRNA was also significantly greater at 12 weeks in laser-dissected highly inflamed areas of the Sgcb-null quadriceps compared to the mdx quadriceps. TGF-β1 was present in areas of degeneration-regeneration, but levels were highly variable and in general did not differ significantly between the two models and controls. The roles of the various subtypes of macrophages in muscle repair and fibrosis in the two models require further study. The Sgcb-null mouse, which develops early fibrosis

  17. Hoxa10 null animals exhibit reduced platelet biogenesis

    PubMed Central

    Konieczna, Iwona M.; DeLuca, Teresa A.; Eklund, Elizabeth A.; Miller, William M.

    2016-01-01

    Summary The transcription factor HOXA10 is an important regulator of myelopoiesis. Engineered over-expression of Hoxa10 in mice results in a myeloproliferative disorder that progresses to acute myeloid leukaemia (AML) over time, and in humans over-expression is associated with poor outcomes in AML. Here, we report that loss of Hoxa10 expression in mice results in reduced platelet count and platelet production, but does not affect clotting efficiency. About 40% fewer platelets were found in Hoxa10 null animals in comparison to wild type littermates. We found a nearly 50% reduction in the percentage of reticulated platelets in Hoxa10 null mice, suggesting deficient platelet production. Furthermore, Hoxa10 null animals recovered less efficiently from induced thrombocytopenia, supporting our hypothesis of defective platelet production. This also correlated with reduced colony formation potential of stem and progenitor cells seeded in megakaryocyte-enhancing conditions in vitro. Together, our results indicate that HOXA10 is important for megakaryopoiesis and platelet biogenesis. PMID:26847476

  18. Partial rescue of the amelogenin null dental enamel phenotype.

    PubMed

    Li, Yong; Suggs, Cynthia; Wright, J Timothy; Yuan, Zhi-an; Aragon, Melissa; Fong, Hanson; Simmons, Darrin; Daly, Bill; Golub, Ellis E; Harrison, Gerald; Kulkarni, Ashok B; Gibson, Carolyn W

    2008-05-30

    The amelogenins are the most abundant secreted proteins in developing dental enamel. Enamel from amelogenin (Amelx) null mice is hypoplastic and disorganized, similar to that observed in X-linked forms of the human enamel defect amelogenesis imperfecta resulting from amelogenin gene mutations. Both transgenic strains that express the most abundant amelogenin (TgM180) have relatively normal enamel, but strains of mice that express a mutated amelogenin (TgP70T), which leads to amelogenesis imperfecta in humans, have heterogeneous enamel structures. When Amelx null (KO) mice were mated with transgenic mice that produce M180 (TgM180), the resultant TgM180KO offspring showed evidence of rescue in enamel thickness, mineral density, and volume in molar teeth. Rescue was not observed in the molars from the TgP70TKO mice. It was concluded that a single amelogenin protein was able to significantly rescue the KO phenotype and that one amino acid change abrogated this function during development.

  19. Balloon exoplanet nulling interferometer (BENI)

    NASA Astrophysics Data System (ADS)

    Lyon, Richard G.; Clampin, Mark; Woodruff, Robert A.; Vasudevan, Gopal; Ford, Holland; Petro, Larry; Herman, Jay; Rinehart, Stephen; Carpenter, Kenneth; Marzouk, Joe

    2009-08-01

    We evaluate the feasibility of a balloon-borne nulling interferometer to detect and characterize an exosolar planet and the surrounding debris disk. The existing instrument consists of a three-telescope Fizeau imaging interferometer with thre fast steering mirrors and three delay lines operating at 800 Hz for closed-loop control of wavefront errors and fine pointing. A compact visible nulling interferometer would be coupled to the imaging interferometer and in principle, allows deep starlight suppression. Atmospheric simulations of the environment above 100,000 feet show that balloonborne payloads are a possible path towards the direct detection and characterization of a limited set of exoplanets and debris disks. Furthermore, rapid development of lower cost balloon payloads provide a path towards advancement of NASA technology readiness levels for future space-based exoplanet missions. Discussed are the BENI mission and instrument, the balloon environment and the feasibility of such a balloon-borne mission.

  20. Nulling at the Keck Interferometer

    NASA Technical Reports Server (NTRS)

    Colavita, M. Mark; Serabyn, Gene; Wizinowich, Peter L.; Akeson, Rachel L.

    2006-01-01

    The nulling mode of the Keck Interferometer is being commissioned at the Mauna Kea summit. The nuller combines the two Keck telescope apertures in a split-pupil mode to both cancel the on-axis starlight and to coherently detect the residual signal. The nuller, working at 10 um, is tightly integrated with the other interferometer subsystems including the fringe and angle trackers, the delay lines and laser metrology, and the real-time control system. Since first 10 um light in August 2004, the system integration is proceeding with increasing functionality and performance, leading to demonstration of a 100:1 on-sky null in 2005. That level of performance has now been extended to observations with longer coherent integration times. An overview of the overall system is presented, with emphasis on the observing sequence, phasing system, and differences with respect to the V2 system, along with a presentation of some recent engineering data.

  1. Balloon Exoplanet Nulling Interferometer (BENI)

    NASA Technical Reports Server (NTRS)

    Lyon, Richard G.; Clampin, Mark; Woodruff, Robert A.; Vasudevan, Gopal; Ford, Holland; Petro, Larry; Herman, Jay; Rinehart, Stephen; Carpenter, Kenneth; Marzouk, Joe

    2009-01-01

    We evaluate the feasibility of using a balloon-borne nulling interferometer to detect and characterize exosolar planets and debris disks. The existing instrument consists of a 3-telescope Fizeau imaging interferometer with 3 fast steering mirrors and 3 delay lines operating at 800 Hz for closed-loop control of wavefront errors and fine pointing. A compact visible nulling interferometer is under development which when coupled to the imaging interferometer would in-principle allow deep suppression of starlight. We have conducted atmospheric simulations of the environment above 100,000 feet and believe balloons are a feasible path forward towards detection and characterization of a limited set of exoplanets and their debris disks. Herein we will discuss the BENI instrument, the balloon environment and the feasibility of such as mission.

  2. Protection from oxidative and electrophilic stress in the Gsta4-null mouse heart

    PubMed Central

    Beneš, Helen; Vuong, Mai K.; Boerma, Marjan; McElhanon, Kevin E.; Siegel, Eric R.; Singh, Sharda P.

    2013-01-01

    4-hydroxynonenal (4-HNE) mediates many pathological effects of oxidative and electrophilic stress and signals to activate cytoprotective gene expression regulated by NF-E2-related factor 2 (Nrf2). By exhibiting very high levels of 4-HNE-conjugating activity, the murine glutathione transferase alpha 4 (GSTA4-4) helps regulate cellular 4-HNE levels. To examine the role of 4-HNE in vivo, we disrupted the murine Gsta4 gene. Gsta4-null mice exhibited no cardiac phenotype under normal conditions and no difference in cardiac 4-HNE level as compared to wild-type (WT) mice. We hypothesized that the Nrf2 pathway might contribute an important compensatory mechanism to remove excess cardiac 4-HNE in Gsta4-null mice. Cardiac nuclear extracts from Gsta4-null mice exhibited significantly higher Nrf2 binding to antioxidant-response elements (AREs). We also observed responses in critical Nrf2 target gene products: elevated Sod2, Cat, and Akr1b7 mRNA levels and significant increases in both cardiac anti-oxidant and anti-electrophile enzyme activities. Gsta4-null mice were less sensitive and maintained normal cardiac function following chronic doxorubicin (DOX) treatment, known to increase cardiac 4-HNE levels. Hence, in the absence of GSTA4-4 to modulate both physiological and pathological 4-HNE levels, the adaptive Nrf2 pathway may be primed to contribute to a preconditioned cardiac phenotype in the Gsta4-null mouse. PMID:23690225

  3. Altitude Differentiated Aerosol Extinction Over Tenerife (North Atlantic Coast) During ACE-2 by Means of Ground and Airborne Photometry and Lidar Measurements

    NASA Technical Reports Server (NTRS)

    Formenti, P.; Elias, T.; Welton, J.; Diaz, J. P.; Exposito, F.; Schmid, B.; Powell, D.; Holben, B. N.; Smirnov, A.; Andreae, M. O.; Devaux, C.; Voss, K.; Lelieveld, J.; Livingston, J. M.; Russell, P. B.; Durkee, P. A.

    2000-01-01

    Retrievals of spectral aerosol optical depths (tau(sub a)) by means of sun photometers have been undertaken in Tenerife (28 deg 16' N, 16 deg 36' W) during ACE-2 (June-July 1997). Five ground-based sites were located at four different altitudes in the marine boundary layer and in the free troposphere, from 0 to 3570 m asl. The goal of the investigation was to provide estimates of the vertical aerosol extinction over the island, both under clean and turbid conditions. Inversion of spectral tau(sub a) allowed to retrieve size distributions, from which the single scattering albedo omega(sub 0) and the asymmetry factor g could be estimated as a function of altitude. These parameters were combined to calculate aerosol forcing in the column. Emphasis is put on episodes of increased turbidity, which were observed at different locations simultaneously, and attributed to outbreaks of mineral dust from North Africa. Differentiation of tau(sub a) as a function of altitude provided the vertical profile of the extinction coefficient sigma(sub e). For dust outbreaks, aerosol extinction is concentrated in two distinct layers above and below the strong subsidence inversion around 1200 m asl. Vertical profiles of tau(sub a) and sigma(sub e) are shown for July 8. In some occasions, vertical profiles are compared to LIDAR observations, performed both at sea level and in the low free troposphere, and to airborne measurements of aerosol optical depths.

  4. RNAi of ace1 and ace2 in Blattella germanica reveals their differential contribution to acetylcholinesterase activity and sensitivity to insecticides.

    PubMed

    Revuelta, L; Piulachs, M D; Bellés, X; Castañera, P; Ortego, F; Díaz-Ruíz, J R; Hernández-Crespo, P; Tenllado, F

    2009-12-01

    Cyclorrhapha insect genomes contain a single acetylcholinesterase (AChE) gene while other insects contain at least two ace genes (ace1 and ace2). In this study we tested the hypothesis that the two ace paralogous from Blattella germanica have different contributions to AChE activity, using RNA interference (RNAi) to knockdown each one individually. Paralogous-specific depletion of Bgace transcripts was evident in ganglia of injected cockroaches, although the effects at the protein level were less pronounced. Using spectrophotometric and zymogram measurements, we obtained evidence that BgAChE1 represents 65-75% of the total AChE activity in nerve tissue demonstrating that ace1 encodes a predominant AChE. A significant increase in sensitivity of Bgace1-interfered cockroaches was observed after 48 h of exposure to chlorpyrifos. In contrast, Bgace2 knockdown had a negligible effect on mortality to this organophosphate. These results point out a key role, qualitative and/or quantitative, of AChE1 as target of organophosphate insecticides in this species. Silencing the expression of Bgace1 but not Bgace2 also produced an increased mortality in insects when synergized with lambda-cyhalothrin, a situation which resembles the synergistic effects observed between organophosphates and pyrethroids. Gene silencing of ace genes by RNAi offers an exciting approach for examining a possible functional differentiation in ace paralogous.

  5. Visualization of the human CD4{sup +} T-cell response in humanized HLA-DR4-expressing NOD/Shi-scid/γc{sup null} (NOG) mice by retrogenic expression of the human TCR gene

    SciTech Connect

    Takahashi, Takeshi Katano, Ikumi; Ito, Ryoji; Ito, Mamoru

    2015-01-02

    Highlights: • β-Lactoglobulin (BLG) specific TCR genes were introduced to human HSC by retrovirus. • Human HSC with BLG-specific TCR were transplanted into NOG-HLA-DR4 I-A{sup −/−} mice. • BLG-specific TCR induced positive selection of thymocytes. • BLG-specific TCR positive CD4{sup +} T cells mediated immune responses in humanized mice. - Abstract: The development of severe immunodeficient mouse strains containing various human genes, including cytokines or HLA, has enabled the reconstitution of functional human immune systems after transplantation of human hematopoietic stem cells (HSC). Accumulating evidence has suggested that HLA-restricted antigen-specific human T-cell responses can be generated in these humanized mice. To directly monitor immune responses of human CD4{sup +} T cells, we introduced β-lactoglobulin (BLG)-specific T cell receptor (TCR) genes derived from CD4{sup +} T-cell clones of cow-milk allergy patients into HSCs, and subsequently transplanted them into NOG-HLA-DR4 transgenic/I-Aβ deficient mice (NOG-DR4/I-A{sup o}). In the thymus, thymocytes with BLG-specific TCR preferentially differentiated into CD4{sup +}CD8{sup −} single-positive cells. Adoptive transfer of mature CD4{sup +} T cells expressing the TCR into recipient NOG-DR4/I-A{sup o} mice demonstrated that human CD4{sup +} T cells proliferated in response to antigenic stimulation and produced IFN-γ in vivo, suggesting that functional T-cell reactions (especially Th1-skewed responses) were induced in humanized mice.

  6. Broken chiral symmetry on a null plane

    SciTech Connect

    Beane, Silas R.

    2013-10-15

    On a null-plane (light-front), all effects of spontaneous chiral symmetry breaking are contained in the three Hamiltonians (dynamical Poincaré generators), while the vacuum state is a chiral invariant. This property is used to give a general proof of Goldstone’s theorem on a null-plane. Focusing on null-plane QCD with N degenerate flavors of light quarks, the chiral-symmetry breaking Hamiltonians are obtained, and the role of vacuum condensates is clarified. In particular, the null-plane Gell-Mann–Oakes–Renner formula is derived, and a general prescription is given for mapping all chiral-symmetry breaking QCD condensates to chiral-symmetry conserving null-plane QCD condensates. The utility of the null-plane description lies in the operator algebra that mixes the null-plane Hamiltonians and the chiral symmetry charges. It is demonstrated that in a certain non-trivial limit, the null-plane operator algebra reduces to the symmetry group SU(2N) of the constituent quark model. -- Highlights: •A proof (the first) of Goldstone’s theorem on a null-plane is given. •The puzzle of chiral-symmetry breaking condensates on a null-plane is solved. •The emergence of spin-flavor symmetries in null-plane QCD is demonstrated.

  7. Hypersensitivities for acetaldehyde and other agents among cancer cells null for clinically relevant Fanconi anemia genes.

    PubMed

    Ghosh, Soma; Sur, Surojit; Yerram, Sashidhar R; Rago, Carlo; Bhunia, Anil K; Hossain, M Zulfiquer; Paun, Bogdan C; Ren, Yunzhao R; Iacobuzio-Donahue, Christine A; Azad, Nilofer A; Kern, Scott E

    2014-01-01

    Large-magnitude numerical distinctions (>10-fold) among drug responses of genetically contrasting cancers were crucial for guiding the development of some targeted therapies. Similar strategies brought epidemiological clues and prevention goals for genetic diseases. Such numerical guides, however, were incomplete or low magnitude for Fanconi anemia pathway (FANC) gene mutations relevant to cancer in FANC-mutation carriers (heterozygotes). We generated a four-gene FANC-null cancer panel, including the engineering of new PALB2/FANCN-null cancer cells by homologous recombination. A characteristic matching of FANCC-null, FANCG-null, BRCA2/FANCD1-null, and PALB2/FANCN-null phenotypes was confirmed by uniform tumor regression on single-dose cross-linker therapy in mice and by shared chemical hypersensitivities to various inter-strand cross-linking agents and γ-radiation in vitro. Some compounds, however, had contrasting magnitudes of sensitivity; a strikingly high (19- to 22-fold) hypersensitivity was seen among PALB2-null and BRCA2-null cells for the ethanol metabolite, acetaldehyde, associated with widespread chromosomal breakage at a concentration not producing breaks in parental cells. Because FANC-defective cancer cells can share or differ in their chemical sensitivities, patterns of selective hypersensitivity hold implications for the evolutionary understanding of this pathway. Clinical decisions for cancer-relevant prevention and management of FANC-mutation carriers could be modified by expanded studies of high-magnitude sensitivities.

  8. Visualization of the human CD4⁺ T-cell response in humanized HLA-DR4-expressing NOD/Shi-scid/γc(null) (NOG) mice by retrogenic expression of the human TCR gene.

    PubMed

    Takahashi, Takeshi; Katano, Ikumi; Ito, Ryoji; Ito, Mamoru

    2015-01-02

    The development of severe immunodeficient mouse strains containing various human genes, including cytokines or HLA, has enabled the reconstitution of functional human immune systems after transplantation of human hematopoietic stem cells (HSC). Accumulating evidence has suggested that HLA-restricted antigen-specific human T-cell responses can be generated in these humanized mice. To directly monitor immune responses of human CD4(+) T cells, we introduced β-lactoglobulin (BLG)-specific T cell receptor (TCR) genes derived from CD4(+) T-cell clones of cow-milk allergy patients into HSCs, and subsequently transplanted them into NOG-HLA-DR4 transgenic/I-Aβ deficient mice (NOG-DR4/I-A(o)). In the thymus, thymocytes with BLG-specific TCR preferentially differentiated into CD4(+)CD8(-) single-positive cells. Adoptive transfer of mature CD4(+) T cells expressing the TCR into recipient NOG-DR4/I-A(o) mice demonstrated that human CD4(+) T cells proliferated in response to antigenic stimulation and produced IFN-γ in vivo, suggesting that functional T-cell reactions (especially Th1-skewed responses) were induced in humanized mice.

  9. Collagen VI deficiency reduces muscle pathology, but does not improve muscle function, in the γ-sarcoglycan-null mouse.

    PubMed

    de Greef, Jessica C; Hamlyn, Rebecca; Jensen, Braden S; O'Campo Landa, Raul; Levy, Jennifer R; Kobuke, Kazuhiro; Campbell, Kevin P

    2016-04-01

    Muscular dystrophy is characterized by progressive skeletal muscle weakness and dystrophic muscle exhibits degeneration and regeneration of muscle cells, inflammation and fibrosis. Skeletal muscle fibrosis is an excessive deposition of components of the extracellular matrix including an accumulation of Collagen VI. We hypothesized that a reduction of Collagen VI in a muscular dystrophy model that presents with fibrosis would result in reduced muscle pathology and improved muscle function. To test this hypothesis, we crossed γ-sarcoglycan-null mice, a model of limb-girdle muscular dystrophy type 2C, with a Col6a2-deficient mouse model. We found that the resulting γ-sarcoglycan-null/Col6a2Δex5 mice indeed exhibit reduced muscle pathology compared with γ-sarcoglycan-null mice. Specifically, fewer muscle fibers are degenerating, fiber size varies less, Evans blue dye uptake is reduced and serum creatine kinase levels are lower. Surprisingly, in spite of this reduction in muscle pathology, muscle function is not significantly improved. In fact, grip strength and maximum isometric tetanic force are even lower in γ-sarcoglycan-null/Col6a2Δex5 mice than in γ-sarcoglycan-null mice. In conclusion, our results reveal that Collagen VI-mediated fibrosis contributes to skeletal muscle pathology in γ-sarcoglycan-null mice. Importantly, however, our data also demonstrate that a reduction in skeletal muscle pathology does not necessarily lead to an improvement of skeletal muscle function, and this should be considered in future translational studies.

  10. Disruptions of Sleep/Wake Patterns in the Stable Tubule Only Polypeptide (STOP) Null Mouse Model of Schizophrenia.

    PubMed

    Profitt, Maxine F; Deurveilher, Samuel; Robertson, George S; Rusak, Benjamin; Semba, Kazue

    2016-09-01

    Disruption of sleep/wake cycles is common in patients with schizophrenia and correlates with cognitive and affective abnormalities. Mice deficient in stable tubule only polypeptide (STOP) show cognitive, behavioral, and neurobiological deficits that resemble those seen in patients with schizophrenia, but little is known about their sleep phenotype. We characterized baseline sleep/wake patterns and recovery sleep following sleep deprivation in STOP null mice. Polysomnography was conducted in adult male STOP null and wild-type (WT) mice under a 12:12 hours light:dark cycle before, during, and after 6 hours of sleep deprivation during the light phase. At baseline, STOP null mice spent more time awake and less time in non-rapid eye movement sleep (NREMS) over a 24-hour period, with more frequent transitions between wake and NREMS, compared to WT mice, especially during the dark phase. The distributions of wake, NREMS and REMS across the light and the dark phases differed by genotype, and so did features of the electroencephalogram (EEG). Following sleep deprivation, both genotypes showed homeostatic increases in sleep duration, with no significant genotype differences in the initial compensatory increase in sleep intensity (EEG delta power). These results indicate that STOP null mice sleep less overall, and their sleep and wake periods are more fragmented than those of WT mice. These features in STOP null mice are consistent with the sleep patterns observed in patients with schizophrenia.

  11. Visible Nulling Coronagraph Testbed Results

    NASA Technical Reports Server (NTRS)

    Lyon, Richard G.; Clampin, Mark; Melnick, Gary; Tolls, Volker; Woodruff, Robert; Vasudevan, Gopal; Rizzo, Maxime; Thompson, Patrick

    2009-01-01

    The Extrasolar Planetary Imaging Coronagraph (EPIC) is a NASA Astrophysics Strategic Mission Concept study and a proposed NASA Discovery mission to image and characterize extrasolar giant planets in orbits with semi-major axes between 2 and 10 AU. EPIC would provide insights into the physical nature of a variety of planets in other solar systems complimenting radial velocity (RV) and astrometric planet searches. It will detect and characterize the atmospheres of planets identified by radial velocity surveys, determine orbital inclinations and masses, characterize the atmospheres around A and F stars, observed the inner spatial structure and colors of inner Spitzer selected debris disks. EPIC would be launched to heliocentric Earth trailing drift-away orbit, with a 5-year mission lifetime. The starlight suppression approach consists of a visible nulling coronagraph (VNC) that enables starlight suppression in broadband light from 480-960 nm. To demonstrate the VNC approach and advance it's technology readiness we have developed a laboratory VNC and have demonstrated white light nulling. We will discuss our ongoing VNC work and show the latest results from the VNC testbed.

  12. Shipboard Sunphotometer Measurements of Aerosol Optical Depth Spectra and Columnar Water Vapor During ACE-2, and Comparison with Selected Land, Ship, Aircraft, and Satellite Measurements

    NASA Technical Reports Server (NTRS)

    Livingston, John M.; Kapustin, Vladimir N.; Schmid, Beat; Russell, Philip B.; Quinn, Patricia K.; Bates, Timothy S.; Durkee, Philip A.; Smith, Peter J.; Freudenthaler, Volker; Wiegner, Matthias

    2000-01-01

    Analyses of aerosol optical depth (AOD) and columnar water vapor (CWV) measurements acquired with NASA Ames Research Center's six-channel Airborne Tracking Sunphotometer (AATS-6) operated aboard the R/V (research vehicle) Professor Vodyanitskiy during the second Aerosol Characterization Experiment (ACE-2) are discussed. Data are compared with various in situ and remote measurements for selected cases. The focus is on 10 July, when the Pelican airplane flew within 70 km of the ship near the time of a NOAA (National Oceanographic and Atmospheric Administration)-14/AVHRR (Advanced Very High Resolution Radiometer) satellite overpass and AOD measurements with the 14-channel Ames Airborne Tracking Sunphotometer (AATS-14) above the marine boundary layer (MBL) permitted calculation of AOD within the MBL from the AATS-6 measurements. A detailed column closure test is performed for MBL AOD on 10 July by comparing the AATS-6 MBL AODs with corresponding values calculated by combining shipboard particle size distribution measurements with models of hygroscopic growth and radiosonde humidity profiles (plus assumptions on the vertical profile of the dry particle size distribution and composition). Large differences (30-80% in the mid-visible) between measured and reconstructed AODs are obtained, in large part because of the high sensitivity of the closure methodology to hygroscopic growth models, which vary considerably and have not been validated over the necessary range of particle size/composition distributions. The wavelength dependence of AATS-6 AODs is compared with the corresponding dependence of aerosol extinction calculated from shipboard measurements of aerosol size distribution and of total scattering measured by a shipboard integrating nephelometer for several days. Results are highly variable, illustrating further the great difficulty of deriving column values from point measurements. AATS-6 CWV values are shown to agree well with corresponding values derived from

  13. Shipboard Sunphotometer Measurements of Aero