Muscle ring finger protein-1 inhibits PKC{epsilon} activation and prevents cardiomyocyte hypertrophy

Much effort has focused on characterizing the signal transduction cascades that are associated with cardiac hypertrophy. In spite of this, we still know little about the mechanisms that inhibit hypertrophic growth. We define a novel anti-hypertrophic signaling pathway regulated by muscle ring finger protein-1 (MURF1) that inhibits the agonist-stimulated PKC-mediated signaling response in neonatal rat ventricular myocytes. MURF1 interacts with receptor for activated protein kinase C (RACK1) and colocalizes with RACK1 after activation with phenylephrine or PMA. Coincident with this agonist-stimulated interaction, MURF1 blocks PKCepsilon translocation to focal adhesions, which is a critical event in the hypertrophic signaling cascade. MURF1 inhibits focal adhesion formation, and the activity of downstream effector ERK1/2 is also inhibited in the presence of MURF1. MURF1 inhibits phenylephrine-induced (but not IGF-1-induced) increases in cell size. These findings establish that MURF1 is a key regulator of the PKC-dependent hypertrophic response and can blunt cardiomyocyte hypertrophy, which may have important implications in the pathophysiology of clinical cardiac hypertrophy.     

A foldable CFTR{Delta}F508 biogenic intermediate accumulates upon inhibition of the Hsc70-CHIP E3 ubiquitin ligase

CFTRDeltaF508 exhibits a correctable protein-folding defect that leads to its misfolding and premature degradation, which is the cause of cystic fibrosis (CF). Herein we report on the characterization of the CFTRDeltaF508 biogenic intermediate that is selected for proteasomal degradation and identification of cellular components that polyubiquitinate CFTRDeltaF508. Nonubiquitinated CFTRDeltaF508 accumulates in a kinetically trapped, but folding competent conformation, that is maintained in a soluble state by cytosolic Hsc70. Ubiquitination of Hsc70-bound CFTRDeltaF508 requires CHIP, a U box containing cytosolic cochaperone. CHIP is demonstrated to function as a scaffold that nucleates the formation of a multisubunit E3 ubiquitin ligase whose reconstituted activity toward CFTR is dependent upon Hdj2, Hsc70, and the E2 UbcH5a. Inactivation of the Hsc70-CHIP E3 leads CFTRDeltaF508 to accumulate in a nonaggregated state, which upon lowering of cell growth temperatures, can fold and reach the cell surface. Inhibition of CFTRDeltaF508 ubiquitination can increase its cell surface expression and may provide an approach to treat CF.     

PDTC, metal chelating compound, induces G1 phase cell cycle arrest in vascular smooth muscle cells through inducing p21Cip1 expression: involvement of p38 mitogen activated protein kinase

Pyrrolidine dithiocarbamate (PDTC), a metal chelating compound, is known to induce cell death in vascular smooth muscle cells (VSMC). However, the molecular mechanism for PDTC-induced VSMC death is not well understood. Addition of PDTC reduced cell growth and DNA synthesis on VSMC in low density conditions. However, in serum depleted medium, PDTC did not affect the cell viability, suggesting that certain factors in serum may mediate the cytotoxic effect of PDTC. Several metal chelators prevented the cell death induced by PDTC. In a serum-deprived condition, addition of exogenous metals, copper, iron, and zinc, restored the cytotoxic effect of PDTC. These data indicate that metals such as copper, iron, and zinc in serum may mediate the cytotoxic effect of PDTC. At low VSMC density in 10% FBS, treatment of PDTC, which induced a cell-cycle block in G1-phase, induced down-regulation of cyclins and CDKs and up-regulation of the CDK inhibitor p21 expression, whereas up-regulation of p27 or p53 by PDTC was not observed. Finally, we determined PDTC-mediated signaling pathway involved in VSMC death. Among relevant pathways, PDTC induced marked activation of p38MAPK and JNK. Expression of dominant negative p38MAPK and SB203580, a p38MAPK specific inhibitor, blocked PDTC-dependent p38MAPK, growth inhibition, and p21 expression. These data demonstrate that the p38MAPK pathway participates in p21 induction, which consequently leads to decrease of cyclin D1/cdk4 and cyclin E/cdk2 complexes and PDTC-dependent VSMC growth inhibition. In conclusion, an understanding of the molecular mechanisms of PDTC in VSMC provides a theoretical basis for clinical approaches using antioxidant therapies in atherosclerosis.     

The Effects of Triiodothyronine on Rat Testis: A Morphometric and Immunohistochemical Study

The aim of present study was to investigate the effects of 3,3′,5-triiodothyronine (T₃) on rat testis both morphometrically and immunohistochemically with determining of insulin-like growth factor I (IGF-I) expression. Adult male Wistar-albino rats used in the study were divided into two groups; control and T₃-treated groups. After T₃ treatment there was observed to be a decrease in testicular weights, diameters of seminiferous tubules and the number of sertoli cells, and an increase in the number of leydig cells (P<0.05). Some of the seminiferous tubule lumens of T₃ administrated rats had cellular debris. IGF-I was localized in sertoli cells, late spermatids and leydig cells of all groups. IGF-I immunoreactivity in T₃ treated rats was higher than in controls in all stages of the cycle of rat seminiferous epithelium, but the staining intensity of leydig cells were similar in both groups. In conclusion, the present results suggest that T₃ may modulate the testicular function by affecting IGF-I activity at the gonadal level.     

Analysis of protein interactions within the cytokinin-signaling pathway of Arabidopsis thaliana

The signal of the plant hormone cytokinin is perceived by membrane-located sensor histidine kinases and transduced by other members of the plant two-component system. In Arabidopsis thaliana, 28 two-component system proteins (phosphotransmitters and response regulators) act downstream of three receptors, transmitting the signal from the membrane to the nucleus and modulating the cellular response. Although the principal signaling mechanism has been elucidated, redundancy in the system has made it difficult to understand which of the many components interact to control the downstream biological processes. Here, we present a large-scale interaction study comprising most members of the Arabidopsis cytokinin signaling pathway. Using the yeast two-hybrid system, we detected 42 new interactions, of which more than 90% were confirmed by in vitro coaffinity purification. There are distinct patterns of interaction between protein families, but only a few interactions between proteins of the same family. An interaction map of this signaling pathway shows the Arabidopsis histidine phosphotransfer proteins as hubs, which interact with members from all other protein families, mostly in a redundant fashion. Domain-mapping experiments revealed the interaction domains of the proteins of this pathway. Analyses of Arabidopsis histidine phosphotransfer protein 5 mutant proteins showed that the presence of the canonical phospho-accepting histidine residue is not required for the interactions. Interaction of A-type response regulators with Arabidopsis histidine phosphotransfer proteins but not with B-type response regulators suggests that their known activity in feedback regulation may be realized by interfering at the level of Arabidopsis histidine phosphotransfer protein-mediated signaling. This study contributes to our understanding of the protein interactions of the cytokinin-signaling system and provides a framework for further functional studies in planta.     

Modulation of β-amyloid precursor protein trafficking and processing by the low density lipoprotein receptor family

Selenium is an essential micronutrient that function through selenoproteins. Selenium deficiency results in lower concentrations of selenium and selenoproteins. The brain maintains it's selenium better than other tissues under low-selenium conditions. Recently, the selenium-containing protein selenoprotein P (Sepp) has been identified as a possible transporter of selenium. The targeted disruption of the selenoprotein P gene (Sepp1) results in decreased brain selenium concentration and neurological dysfunction, unless selenium intake is excessive However, the effect of selenoprotein P deficiency on the processes of memory formation and synaptic plasticity is unknown. In the present studies Sepp1(-/-) mice and wild type littermate controls (Sepp1(+/+)) fed a high-selenium diet (1 mg Se/kg) were used to characterize activity, motor coordination, and anxiety as well as hippocampus-dependent learning and memory. Normal associative learning, but disrupted spatial learning was observed in Sepp1(-/-) mice. In addition, severe alterations were observed in synaptic transmission, short-term plasticity and long-term potentiation in hippocampus area CA1 synapses of Sepp1(-/-) mice on a 1 mg Se/kg diet and Sepp1(+/+) mice fed a selenium-deficient (0 mg Se/kg) diet. Taken together, these data suggest that selenoprotein P is required for normal synaptic function, either through presence of the protein or delivery of required selenium to the CNS.     

Economic evaluation of weight loss interventions in overweight and obese women

Objective: To conduct a clinical and economic evaluation of outpatient weight loss strategies in overweight and obese adult U.S. women. Research Methods and Procedures: This study was a lifetime cost‐use analysis from a societal perspective, using a first‐order Monte Carlo simulation. Strategies included routine primary care and varying combinations of diet, exercise, behavior modification, and/or pharmacotherapy. Primary data were collected to assess program costs and obesity‐related quality of life. Other data were obtained from clinical trials, population‐based surveys, and other published literature. This was a simulated cohort of healthy 35‐year‐old overweight and obese women in the United States. Results: For overweight and obese women, a three‐component intervention of diet, exercise, and behavior modification cost $12,600 per quality‐adjusted life year gained compared with routine care. All other strategies were either less effective and more costly or less effective and less cost‐effective compared with the next best alternative. Results were most influenced by obesity‐related effects on quality of life and the probabilities of weight loss maintenance. Discussion: A multidisciplinary weight loss program consisting of diet, exercise, and behavior modification provides good value for money, but more research is required to confirm the impacts of such programs on quality of life and the likelihood of long‐term weight loss maintenance.     

Squatting facet: a case study Dilkaya and Van-Kalesi populations in eastern Turkey

Anomalies of the skeleton provide information on living conditions, cultural structure and health problems in ancient societies. Squatting facet is a kind of anomaly that forms on the surfaces where the tibia and talus articulate is the squatting facet states the daily activities and living style of the society. The aim of this study is to learn the daily activities of the medieval societies in the Van region through studying of squatting facets. In this study, adult skeletons from Dilkaya and Van Kalesi-Eski Van Sehri societies dating to the Medieval Age were investigated (65 tibia and 82 tali from Dilkaya, 61 tibia and 52 tali from Van Kalesi-Eski Van Sehri). The lateral squatting facet had high ratios in both societies. The tibia lateral squatting facet found on females and males of Dilkaya was 97.2% and 96.9%, respectively, and on females and males of Van kalesi Eski Van Sehri was 87.5% and 89.2%, respectively. The talus lateral squatting facet found on females and males of Dilkaya was 72.1% and 51.3%, respectively, and on females and males of Van kalesi Eski Van Sehri was 91.2% and 83.7%, respectively. The results provide an opportunity to study the relationship between past and modern population, and also describe the daily activity of life and cultural structure.     

Adaptation of rat gastric tissue against indomethacin toxicity

Indomethacin is used in the treatment of inflammatory diseases. But the drug toxicity limits its usage. This study investigated whether adaptation occurred after various dosages of repeated (chronic) indomethacin in rats to the gastro-toxic effects of indomethacin. It also examined whether the adaptation was related to oxidant–antioxidant mechanisms and oxidative DNA damage in gastric tissue. To illuminate the adaptation mechanism in the gastric tissue of rats given various dosages of chronic indomethacin, the levels of oxidants and antioxidants (GSH, MDA, NO, SOD and MPO), activities of COX-1 and COX-2 enzymes and oxidative DNA damage (8-OHd Gua/10⁵ Gua) were measured. Results were compared to 25-mg/kg single-dose indomethacin group, and the role of oxidant and antioxidant parameters and oxidative DNA damage in the adaptation mechanism was evaluated. The average ulcer areas of gastric tissue of the 0.5-, 1-, 2-, 3-, 4-, and 5-mg/kg dosages of chronic indomethacin given to rats were 19.5 ± 3.7, 12.5 ± 3.3, 10 ± 5.2, 4.5 ± 3.6, 8.6 ± 2.4, and 9.5 ± 2.1 mm², respectively. This rate was measured as 21.3 ± 2.6 mm² in the single-dose indomethacin group. Consequently, after various dosages of repeated (chronic) indomethacin administration in rats, it was observed that a clear adaptation developed against gastric damage and that gastric damage was reduced. The best adaptation was observed in the gastric tissue of the 3-mg/kg chronic indomethacin group. In parallel with the damage reduction, the oxidant parameters (MDA and MPO) and oxidative DNA damage (8-OHd Gua/10⁵ Gua) were reduced, and the antioxidant parameters (GSH, NO and SOD) were increased. There is no relation between COX enzymes and adaptation mechanism. This circumstance shows that not COX-1 and COX-2 enzymes, oxidant and antioxidant parameters may play a role in the adaptation mechanism.     

ATP stimulates MMP-2 release from human aortic smooth muscle cells via JNK signaling pathway

Aortic smooth muscle cell release of matrix metalloproteinase-2 (MMP-2) and tissue inhibitor of metalloproteinase-2 (TIMP-2) has been implicated in aortic aneurysm pathogenesis, but proximal modulation of release is poorly understood. Extracellular nucleotides regulate vascular smooth muscle cell metabolism in response to physiochemical stresses, but nucleotide modulation of MMP and/or TIMP release has not been reported. We hypothesized that nucleotides modulate MMP-2 and TIMP-2 release from human aortic smooth muscle cells (HASMCs) via distinct purinergic receptors and signaling pathways. We exposed HASMCs to exogenous ATP and other nucleotides with and without interleukin-1beta (IL-1beta). HASMCs were pretreated in some experiments with apyrase, which degrades ATP, and inhibitors of ERK1/2, JNK, and p38 MAPK. MMP-2 and TIMP-2 released into supernatant were assessed using ELISA and Western blotting. ATP, adenosine, and UTP significantly stimulated MMP-2 release in the presence of IL-1beta (300 nM ATP: 181 +/- 22%, P = 0.003; 30 microm adenosine: 244 +/- 150%, P = 0.001; and 200 microm UTP: 153 +/- 40%, P = 0.015; vs. 100% constitutive). ATP also stimulated MMP-2 release in the absence of IL-1beta (100 microm ATP: 148 +/- 38% vs. 100% constitutive). Apyrase significantly reduced ATP-stimulated MMP-2 release (apyrase + 500 nM ATP: 59 +/- 3% vs. 124 +/- 7% with 500 nM ATP). Rank-order agonist potency for MMP-2 release was consistent with ATP activation of PAY and PAY receptors. ATP induced phosphorylation of intracellular JNK, and inhibition of the JNK pathway blocked ATP-stimulated MMP-2 release, indicating signaling via this pathway. Nucleotides are thus novel stimulants of MMP-2 release from HASMCs and may provide a mechanistic link between physiochemical stress in the aorta and aneurysms, especially in the context of inflammation.