Ubiquitin-Independent Proteasomal Degradation Mediated by Antizyme

Most of the proteins in eukaryotic cells are degraded by the proteasome in an ubiquitin-dependent manner. However, ubiquitin-independent protein degradation pathway by the 26S proteasome exists in the cells. Ornithine decarboxylase (ODC) is a well-known protein that is degraded by the 26S proteasome without ubiquitination. Degradation of ODC requires the protein, “antizyme (AZ),” that is induced by polyamine and binds to the ODC monomer to inhibit ODC activity and target it to the 26S proteasome for proteolytic degradation. Namely, AZ contributes the feedback regulation of intracellular polyamine level. ODC has been considered to be the only protein that AZ binds and accelerates its degradation. However, recently AZ-mediated proteasomal protein degradation will gradually increase. Most recently, we found that one of the antizyme families, AZ2, accelerates c-Myc degradation by the proteasome without ubiquitination. In this chapter, we introduce latest several ubiquitin-independent proteasomal degradation mediated by antizyme

Conditional deletion of Bmpr1a in differentiated osteoclasts increases osteoblastic bone formation, increasing volume of remodeling bone in mice

Bone undergoes remodeling consisting of osteoclastic bone resorption followed by osteoblastic bone formation throughout life. Although the effects of bone morphogenetic protein (BMP) signals on osteoblasts have been studied extensively, the function of BMP signals in osteoclasts has not been fully elucidated. To delineate the function of BMP signals in osteoclasts during bone remodeling, we deleted BMP receptor type IA ( Bmpr1a ) in an osteoclast‐specific manner using a knock‐in Cre mouse line to the cathepsin K locus ( Ctsk Cre/+ ;Bmpr1a flox/flox , designated as Bmpr1a ΔOc/ΔOc ). Cre was specifically expressed in multinucleated osteoclasts in vivo. Cre‐dependent deletion of the Bmpr1a gene occurred at 4 days after cultivation of bone marrow macrophages obtained from Bmpr1a ΔOc/ΔOc with RANKL. These results suggested that Bmpr1a was deleted after formation of osteoclasts in Bmpr1a ΔOc/ΔOc mice. Expression of bone‐resorption markers increased, thus suggesting that BMPRIA signaling negatively regulates osteoclast differentiation. Trabeculae in tibia and femurs were thickened in 3.5‐, 8‐, and 12‐week‐old Bmpr1a ΔOc/ΔOc mice. Bone histomorphometry revealed increased bone volume associated with increased osteoblastic bone‐formation rates (BFR) in the remodeling bone of the secondary spongiosa in Bmpr1a ΔOc/ΔOc tibias at 8 weeks of age. For comparison, we also induced an osteoblast‐specific deletion of Bmpr1a using Col1a1‐Cre. The resulting mice showed increased bone volume with marked decreases in BFR in tibias at 8 weeks of age. These results indicate that deletion of Bmpr1a in differentiated osteoclasts increases osteoblastic bone formation, thus suggesting that BMPR1A signaling in osteoclasts regulates coupling to osteoblasts by reducing bone‐formation activity during bone remodeling. © 2011 American Society for Bone and Mineral ResearchPeer Reviewedhttp://deepblue.lib.umich.edu/bitstream/2027.42/87086/1/477_ftp.pdfhttp://deepblue.lib.umich.edu/bitstream/2027.42/87086/2/jbmr_477_sm_SupplData.pd

Comparison study between the mechanisms of allergic asthma amelioration by a cysteinylleukotriene type 1 receptor antagonist montelukast and methylprednisolone

ABSTRACT We investigated the effects of cysteinyl-leukotriene (cysLT) type 1 receptor antagonist montelukast (MK) and compared them with those of methylprednisolone (MP) in an allergic asthma model. Rats sensitized to ovalbumin (OVA) received repeated intratracheal exposure to OVA for up to 3 consecutive days. Pretreatment with MK or MP before OVA exposure inhibited late airway response (LAR) and reduced cellular infiltration into the bronchial submucosa after the triple OVA. The amount of N-acetyl-leukotriene E 4 in the bile was significantly reduced by pretreatment with MK or MP, suggesting that both drugs reduced the production of cysLTs in the lungs. In the in vitro study, when the fragments of lungs that had been repeatedly pretreated with MK or MP and exposed to OVA were removed and incubated with OVA, the coaddition of either drug significantly reduced cysLT production. In contrast, the cysLT production following the addition of OVA to the lung fragments that had not received in vivo pretreatment with either drug was inhibited by MK but not by MP. These results indicate that MK and MP inhibit LAR by suppressing the infiltration of inflammatory cells into the bronchial submucosa, thereby inhibiting the production of cysLTs in the lungs, and that MK but not MP may inhibit cysLT production directly. The different effects on cysLT production between the two drugs may provide a rationale for the use of combination therapy with cysLT 1 receptor antagonists and steroids for the treatment of asthma

Smad6/Smurf1 overexpression in cartilage delays chondrocyte hypertrophy and causes dwarfism with osteopenia

Biochemical experiments have shown that Smad6 and Smad ubiquitin regulatory factor 1 (Smurf1) block the signal transduction of bone morphogenetic proteins (BMPs). However, their in vivo functions are largely unknown. Here, we generated transgenic mice overexpressing Smad6 in chondrocytes. Smad6 transgenic mice showed postnatal dwarfism with osteopenia and inhibition of Smad1/5/8 phosphorylation in chondrocytes. Endochondral ossification during development in these mice was associated with almost normal chondrocyte proliferation, significantly delayed chondrocyte hypertrophy, and thin trabecular bone. The reduced population of hypertrophic chondrocytes after birth seemed to be related to impaired bone growth and formation. Organ culture of cartilage rudiments showed that chondrocyte hypertrophy induced by BMP2 was inhibited in cartilage prepared from Smad6 transgenic mice. We then generated transgenic mice overexpressing Smurf1 in chondrocytes. Abnormalities were undetectable in Smurf1 transgenic mice. Mating Smad6 and Smurf1 transgenic mice produced double-transgenic pups with more delayed endochondral ossification than Smad6 transgenic mice. These results provided evidence that Smurf1 supports Smad6 function in vivo

A Distinct Structure Inside the Galactic Bar

We present the result of a near-infrared (J H Ks) survey along the Galactic plane, -10.5deg < l < +10.5deg and b=+1.0deg, with the IRSF 1.4m telescope and the SIRIUS camera. Ks vs. H-Ks color-magnitude diagrams reveal a well-defined population of red clump (RC) stars whose apparent magnitude peak changes continuously along the Galactic plane, from Ks=13.4 at l=-10deg to Ks=12.2 at l=+10deg after dereddening. This variation can be explained by the bar-like structure found in previous studies, but we find an additional inner structure at |l| < 4deg, where the longitude - apparent magnitude relation is distinct from the outer bar, and the apparent magnitude peak changes by only 0.1 mag over the central 8deg. The exact nature of this inner structure is as yet uncertain.Comment: 8 pages, 4 figures. accepted by ApJ

Indocyanine Green (ICG) Lymphography Is Superior to Lymphoscintigraphy for Diagnostic Imaging of Early Lymphedema of the Upper Limbs

BACKGROUND: Secondary lymphedema causes swelling in limbs due to lymph retention following lymph node dissection in cancer therapy. Initiation of treatment soon after appearance of edema is very important, but there is no method for early diagnosis of lymphedema. In this study, we compared the utility of four diagnostic imaging methods: magnetic resonance imaging (MRI), computed tomography (CT), lymphoscintigraphy, and Indocyanine Green (ICG) lymphography. PATIENTS AND METHODS: Between April 2010 and November 2011, we examined 21 female patients (42 arms) with unilateral mild upper limb lymphedema using the four methods. The mean age of the patients was 60.4 years old (35-81 years old). Biopsies of skin and collecting lymphatic vessels were performed in 7 patients who underwent lymphaticovenous anastomosis. RESULTS: The specificity was 1 for all four methods. The sensitivity was 1 in ICG lymphography and MRI, 0.62 in lymphoscintigraphy, and 0.33 in CT. These results show that MRI and ICG lymphography are superior to lymphoscintigraphy or CT for diagnosis of lymphedema. In some cases, biopsy findings suggested abnormalities in skin and lymphatic vessels for which lymphoscintigraphy showed no abnormal findings. ICG lymphography showed a dermal backflow pattern in these cases. CONCLUSIONS: Our findings suggest the importance of dual diagnosis by examination of the lymphatic system using ICG lymphography and evaluation of edema in subcutaneous fat tissue using MRI

Early Detection of Abnormal Prion Protein in Genetic Human Prion Diseases Now Possible Using Real-Time QUIC Assay

Introduction: The definitive diagnosis of genetic prion diseases (gPrD) requires pathological confirmation. To date, diagnosis has relied upon the finding of the biomarkers 14-3-3 protein and total tau (t-tau) protein in the cerebrospinal fluid (CSF), but many researchers have reported that these markers are not sufficiently elevated in gPrD, especially in Gerstmann-Sträussler-Scheinker syndrome (GSS). We recently developed a new in vitro amplification technology, designated "real-time quaking-induced conversion (RT-QUIC)", to detect the abnormal form of prion protein in CSF from sporadic Creutzfeldt-Jakob disease (sCJD) patients. In the present study, we aimed to investigate the presence of biomarkers and evaluate RT-QUIC assay in patients with gPrD, as the utility of RT-QUIC as a diagnostic tool in gPrD has yet to be determined. Method/Principal Findings: 56 CSF samples were obtained from gPrD patients, including 20 cases of GSS with P102L mutation, 12 cases of fatal familial insomnia (FFI; D178N), and 24 cases of genetic CJD (gCJD), comprising 22 cases with E200K mutation and 2 with V203I mutation. We subjected all CSF samples to RT-QUIC assay, analyzed 14-3-3 protein by Western blotting, and measured t-tau protein using an ELISA kit. The detection sensitivities of RT-QUIC were as follows: GSS (78%), FFI (100%), gCJD E200K (87%), and gCJD V203I (100%). On the other hand the detection sensitivities of biomarkers were considerably lower: GSS (11%), FFI (0%), gCJD E200K (73%), and gCJD V203I (67%). Thus, RT-QUIC had a much higher detection sensitivity compared with testing for biomarkers, especially in patients with GSS and FFI. Conclusion/Significance: RT-QUIC assay is more sensitive than testing for biomarkers in gPrD patients. RT-QUIC method would thus be useful as a diagnostic tool when the patient or the patient\u27s family does not agree to genetic testing, or to confirm the diagnosis in the presence of a positive result for genetic testing

Feasibility of methotrexate discontinuation following tocilizumab and methotrexate combination therapy in patients with long-standing and advanced rheumatoid arthritis: a 3-year observational cohort study

Objectives: Methotrexate (MTX) is associated with extensive side effects, including myelosuppression, interstitial pneumonia, and infection. It is, therefore, critical to establish whether its administration is required after achieving remission with tocilizumab (TCZ) and MTX combination therapy in patients with rheumatoid arthritis (RA). Therefore, the aim of this multicenter, observational, cohort study was to evaluate the feasibility of MTX discontinuation for the safety of these patients. Methods: Patients with RA were administered TCZ, with or without MTX, for 3 years; those who received TCZ+MTX combination therapy were selected. After remission was achieved, MTX was discontinued without flare development in one group (discontinued [DISC] group, n = 33) and continued without flare development in another group (maintain [MAIN] group, n = 37). The clinical efficacy of TCZ+MTX therapy, patient background characteristics, and adverse events were compared between groups. Results: The disease activity score in 28 joints-erythrocyte sedimentation rate (DAS28-ESR) at 3, 6, and 9 months was significantly lower in the DISC group (P < .05, P < .01, and P < .01, respectively). Further, the DAS28-ESR remission rate at 6 and 9 months and Boolean remission rate at 6 months were significantly higher in the DISC group (P < .01 for all). Disease duration was significantly longer in the DISC group (P < .05). Furthermore, the number of patients with stage 4 RA was significantly higher in the DISC group (P < .01). Conclusions: Once remission was achieved, MTX was discontinued in patients who responded favorably to TCZ+MTX therapy, despite the prolonged disease duration and stage progression

A Comparison of Partial and Full Median Sternotomy for Re-do Cardiac Valve Surgery

Background. Minimally invasive cardiac surgery (MICS) has been reported to reduce surgical trauma, postoperative pain, blood loss, and length of stay. This study compares MICS using partial sternomy with full sternotomy in re- do cardiac valve surgery. Methods. The records of 20 patients who underwent full median sternotomy (group F) and 17 patients who underwent MICS (group M) for re- do cardiac valve surgery from April 1990 to April 2001 were compared retrospectively. Results. Time of operation (skin-to-skin), perfusion time, time of extubation, and length of Intensive Care Unit stay were shorter in group M than group F (352.19 ± 76.05 min vs. 510.70 ± 256.26 min, 143.65 ± 29.41 min vs. 254.60 ± 192.72 min, 16.31 ± 7.56 h vs. 48.47 ± 40.14 h, 2.13 ± 0.81 days vs. 4.53 ± 2.20 days). Intra operative blood loss and chest drainage also were less in group M than group F (774.53 ± 415.48 mL vs. 3781.30 ± 5207.07 mL, 779.70 ± 666.48 mL vs. 1687.56 ± 984.23 mL). Conclusions. This study demonstrates the advantages of MICS for re-do cardiac valve surgery