bluuebunny/crossref_metadata_2025 huggingface dataset:
Era and RbfA Have Overlapping Function in Ribosome Biogenesis in Escherichia coli
Authors: Inoue Koichi , Chen Jingqiu , Tan Qian , Inouye Masayori
Published: July 2006

A cold-shock protein, RbfA (ribosome-binding factor A), is essential for cell growth at low temperature. In an rbfA-deletion strain, 30S and 50S ribosomal subunits increase relative to 70S monosomes with concomitant accumulation of a precursor 16S rRNA (17S rRNA). Recently, we have reported that overexpression of Era, an essential GTP-binding protein, suppresses not only the cold-sensitive cell growth but also defective ribosome biogenesis in the rbfA-deletion strain. Here, in order to elucidate how RbfA and Era functionally overlap, we characterized a cold-sensitive Era mutant (a point mutation at the Glu-200 to Lys; E200K) which shows a similar phenotype as the rbfA-deletion strain; accumulation of free ribosome subunits and 17S rRNA. To examine the effect of E200K in the rbfA-deletion strain, we constructed an E200K-inducible expression system. Interestingly, unlike wild-type Era, overexpression of Era(E200K) protein in the rbfA-deletion strain severely inhibited cell growth even at permissive temperature with further concomitant reduction of 16S rRNA. Purified Era(E200K) protein binds to 30S ribosomal subunits in a nucleotide-dependent manner like wild-type Era and retains both GTPase and autophosphorylation activities. Furthermore, we isolated spontaneous revertants of the E200K mutant. These revertants partially suppressed the accumulation of 17S rRNA. All the spontaneous mutations were found to result in higher Era(E200K) expression. These results suggest that the Era(E200K) protein has an impaired function in ribosome biogenesis without losing its ribosome binding activity. The severe growth defect caused by E200K in the rbfA-deletion strain may be due to competition between intrinsic wild-type Era and overexpressed Era(E200K) for binding to 30S ribosomal subunits. We propose that Era and RbfA have an overlapping function that is essential for ribosome biogenesis, and that RbfA becomes dispensable only at high temperatures because Era can complement its function only at higher temperatures.
Mass Spectrometric Identification of Proteins in Complex Post-Genomic Projects
Authors: Hufnagel Peter , Rabus Ralf
Published: July 2006

The rapidly developing proteomics technologies help to advance the global understanding of physiological and cellular processes. The lifestyle of a study organism determines the type and complexity of a given proteomic project. The complexity of this study is characterized by a broad collection of pathway-specific subproteomes, reflecting the metabolic versatility as well as the regulatory potential of the aromatic-degrading, denitrifying bacterium ‘Aromatoleum’ sp. strain EbN1. Differences in protein profiles were determined using a gel-based approach. Protein identification was based on a progressive application of MALDI-TOF-MS, MALDI-TOF-MS/MS and LC-ESI-MS/MS. This progression was result-driven and automated by software control. The identification rate was increased by the assembly of a project-specific list of background signals that was used for internal calibration of the MS spectra, and by the combination of two search engines using a dedicated MetaScoring algorithm. In total, intelligent bioinformatics could increase the identification yield from 53 to 70% of the analyzed 5,050 gel spots; a total of 556 different proteins were identified. MS identification was highly reproducible: most proteins were identified more than twice from parallel 2DE gels with an average sequence coverage of >50% and rather restrictive score thresholds (Mascot ≧95, ProFound ≧2.2, MetaScore ≧97). The MS technologies and bioinformatics tools that were implemented and integrated to handle this complex proteomic project are presented. In addition, we describe the basic principles and current developments of the applied technologies and provide an overview over the current state of microbial proteome research.
The Cecropin Superfamily of Toxic Peptides
Authors: Tamang Dorjee G. , Saier Jr. Milton H.
Published: July 2006

All sequenced peptide toxins of the cecropin, pleurocidin and dermaceptin/ceratotoxin families in the National Center for Biotechnology Information (NCBI) database as of May 2005 were identified and shown to comprise a single superfamily. The peptide sequences were multiply aligned, revealing conserved residues that may play roles in structure and function. Signature sequences were derived for each of the 3 constituent families. Phylogenetic analyses revealed the relationships of these peptides to each other, and average hydropathy/amphipathicity studies provided structural information. This study serves to characterize a large superfamily of toxic peptides that perform host defense functions in a range of animal kingdoms.
Mechanism of CcpA-Mediated Glucose Repression of the resABCDE Operon of Bacillus subtilis
Authors: Choi Soo-Keun , Saier Jr. Milton H.
Published: July 2006

The resABCDE operon of Bacillus subtilis encodes a three-protein complex involved in cytochrome c biogenesis as well as the ResE sensor kinase and the ResD response regulator that control electron transfer and other functions in response to oxygen availability. We have investigated the mechanism of CcpA-mediated control of res operon expression which occurs maximally in the stationary phase of growth. Two CcpA-binding (CRE) sites were found in the res operon, one (CRE1) in the control region in front of the resA promoter, the other (CRE2) in the resB structural gene. Both CRE sites proved to be essential for full CcpA-mediated glucose repression of res operon expression. We propose that both looping and road block mechanisms are involved in res operon control by CcpA.
Is HCl Duloxetine Effective in the Management of Urinary Stress Incontinence after Radical Prostatectomy?
Authors: Zahariou Athanasios , Papaioannou Polyanthi , Kalogirou Georgia
Published: July 2006

Introduction: Up to 70% of patients who undergo radical prostatectomy complain about urine leakage, but persistent stress incontinence 1 year after surgery affects <5% of them. HCl duloxetine is a dual serotonin and norepinephrine reuptake inhibitor that relieves the symptoms of stress urinary incontinence. The purpose of this study was to evaluate the efficacy of HCl duloxetine in the management of urinary incontinence after radical prostatectomy and its impact in urodynamic parameters such as maximal urethral closure pressure (MUCP), abdominal leak point pressure (ALPP) and retrograde leak point pressure (RLPP). Material and Methods: The study included 18 men with stress urinary incontinence 12 months after radical prostatectomy. All underwent a pad test to quantify the degree of urine loss and a urodynamic evaluation before and after a three month treatment with HCl duloxetine. The intrinsic sphincter was evaluated by ALPP and RLPP and the striated sphincter by MUCP. Results: At the pretreatment evaluation the mean ALPP was 52.1 cm H2O, the mean MUCP was 52.5 cm H2O and the mean RLPP was 43.1 cm H2O. After 3 months of HCl duloxetine treatment the mean ALPP was 59.1 cm H2O, the mean MUCP was 67.3 cm H2O and the mean RLPP was 45.1 cm H2O. There was a statistically significant correlation among RLPP, MUCP and ALPP before treatment. After HCl duloxetine treatment there was significant correlation between RLPP and ALPP. Conclusion: The use of HCl duloxetine results in mild increase of MUCP and in significant reduction of urine loss. Its action on the extrinsic sphincter does not provide a complete treatment option for postprostatectomy incontinence.
Bladder-Sparing Surgery in Locally Advanced Nonurological Pelvic Malignancy
Authors: Siva Prasad G. , Chacko K.N. , Antony D. , Lionel G. , Kekre N.S. , Gopalakrishnan G.
Published: July 2006

Introduction: The urinary bladder is commonly involved in pelvic malignancy. The incidence of apparent extension into adjacent organs in locally advanced colorectal malignancy is 5–12%. It is not known with other pelvic malignancy. No guidelines are available for its management. Often a dilemma exists between cystectomy and a bladder-sparing procedure. We studied the validity of bladder-sparing surgery (BSS) in locally advanced nonurological pelvic malignancy. Methods: Hospital records of patients who underwent BSS along with other surgeries (abdomino-perineal resection, anterior resection, anterior exenteration, debulking surgery and total pelvic exenteration) from January 1992 to May 2003 were reviewed. Results: BSS was done in 15 patients. 10 had locally advanced colorectal malignancy, 3 with soft tissue masses of the lateral pelvic wall, 1 had ovarian malignancy and the other had residual mass following radiotherapy and chemotherapy of cancer cervix. In those with locally advanced colorectal malignancy, symptoms suggestive of lower urinary tract involvement were present in 8 (80%). Urine examination and ultrasonography was not helpful in suggesting bladder involvement, unlike CT scan of abdomen and pelvis. Preoperative cystoscopy showed endoscopic evidence of bladder involvement in 7 (87.5%). Bladder was involved supratrigonally in 7. Partial cystectomy was done in 9 patients. The left ureter was involved in 6 patients, and they required ureteric reimplantation. Palliative transurethral resection was done in 1 patient with tumor infiltration at the bladder neck and prostate. 50% patients had bothersome lower urinary tract symptoms at 1 year. One patient died in the immediate postoperative period due to a nonurological cause. Overall 3-year survival rate was 40%. Conclusion: Unlike primary bladder cancers these lesions are not multifocal and hence en block conservative bladder-sparing surgery can be offered. Preoperative CT scan or MRI can predict lower urinary tract involvement and help in decision-making by both surgeon and patient. The ultimate decision for bladder sparing is based on intraoperative findings. Sparing the bladder might provide better quality of life by avoiding urinary diversion without altering survival.
Adjuvant Chemotherapy in the Management of pT3N0M0 Transitional Cell Carcinoma of the Upper Urinary Tract
Authors: Lee Sang Eun , Byun Seok-Soo , Park Yong Hyun , Chang In Ho , Kim Yong June , Hong Sung Kyu
Published: July 2006

Introduction: We investigate the efficacy of postoperative adjuvant chemotherapy for locally advanced, but lymph node negative, pathologic stage T3 transitional cell carcinoma (TCC) of the upper urinary tract. Patients and Methods: A retrospective study on 27 patients who had undergone radical nephroureterectomy with regional lymphadenectomy for pT3N0M0 primary upper urinary tract TCC at our institution from 1996 to 2001 was performed. Among the 27 patients, 16 also received adjuvant chemotherapy following surgery (adjuvant group), whereas the other 11 patients did not (nonadjuvant group). Results: Adjuvant and nonadjuvant therapy groups were not significantly different with respect to age, sex, performance status, tumor grade, and tumor location. Overall, 5 of the 16 patients (31%) in the adjuvant group and 4 of the 11 patients (36%) in the nonadjuvant group had recurrence of cancer at 40 months of follow-up. The two groups demonstrated no significant differences in recurrence-free survival (p = 0.794) and disease-specific survival (p = 0.783). Conclusions: Although it would be difficult to draw any definite conclusions from the results of our investigations, our data suggest that adjuvant therapy with traditional conventional chemotherapeutic regimens alone may not be effective as previously anticipated in significantly improving survival rates for locally advanced, but lymph node negative, TCC of the upper urinary tract.
Urgent Superselective Segmental Renal Artery Embolization in the Treatment of Life-Threatening Renal Hemorrhage
Authors: Pappas Paris , Leonardou Polytimi , Papadoukakis Stefanos , Zavos George , Michail Spyridon , Boletis John , Tzortzis George
Published: July 2006

Introduction: Renal hemorrhage is a major life-threatening condition that can be caused by trauma, operation, biopsy, as well as sudden spontaneous rupture of renal tumors or aneurysms. We report our experience with superselective segmental renal artery catheterization and embolization as therapeutic options for such cases. Patients and Methods: Over the last 8 years, 28 patients with severe renal hemorrhage were admitted for evaluation and possible further treatment. Twenty of them had a history of previous biopsy (6 of them one of a transplanted kidney), 1 patient had a recent percutaneous nephrostomy, 4 patients presented with renal mass ruptures (2 patients renal cell carcinoma, 1 patient angiomyolipoma, 1 patient hemorrhagic cysts), 1 patient had rupture of a renal aneurysm during delivery, 1 patient suffered bleeding after partial nephrectomy, and 1 patient was hospitalized after a car accident. They all presented with clinical signs of hemodynamic instability. Angiographic investigation of the kidneys preceded further intervention in all cases. 26 out of the 28 patients underwent superselective embolization of the specific bleeding vessel with the use of microcoils and/or Gelfoam particles. Results: All patients treated by superselective segmental renal artery embolization had a successful outcome, including a steady renal function and a stable clinical course. No complications occurred. Conclusion: Superselective segmental renal artery catheterization and embolization is a safe and efficient method for the treatment of patients with severe renal hemorrhage, preserving healthy renal parenchyma and renal function.
Bladder Squamous Metaplasia of the Urothelium – Introductory Report
Authors: Jurkiewicz Beata , Matuszewski Łukasz , Bokwa Tadeusz , Gut Grzegorz , Rybak Dorota
Published: July 2006

Seven female patients hospitalized in the Surgery Ward of Warsaw’s Hospital for Children are described. Nonspecific abdominal pains, recurrent urinary tract infections, and voiding disorders (difficulty in voiding commencement) were the primary causes of hospitalization. Urodynamics and cystoscopy with a histopathology section of the urothelium achieved the diagnosis of nonkeratizing squamous cell metaplasia of the urothelium. The topic is not new, although the diagnosis is particularly rare in the pediatric patient. Until now, the existing world literature has described only a few cases of this kind of metaplasia in children. These findings may be helpful in differential diagnosis of urinary bladder disease. It is postulated that nonkeratizing squamous cell metaplasia of the urothelium, in the future, may lead to malignant changes in the urothelium. Therefore, the therapeutic process merits close monitoring.
Serum Hemoglobin Levels Predict Response to Strontium-89 and Rhenium-186-HEDP Radionuclide Treatment for Painful Osseous Metastases in Prostate Cancer
Authors: van der Poel H.G. , Antonini N. , Hoefnagel C.A. , Horenblas S. , Valdes Olmos R.A.
Published: July 2006

Objective: Retrospective comparative analysis of strontium-89 chloride (Sr89) and rhenium-186-hydroxyethylidene diphosphonate (Re186-HEDP) radionuclide treatment to find predictors of response in patients with painful metastases from hormone refractory prostate cancer. Patients and Methods: Clinical data from 60 hormone refractory PCA patients (i.e. rising PSA at castrate testosterone serum levels) was obtained. Twenty-nine were treated with Sr89, 31 were treated with Re186-HEDP for painful osseous metastasis. Response was defined as a patient-reported decrease in pain and/or reduction in pain medication with stable pain level. Hematological parameters and serum levels of PSA, alkaline phosphatase, and lactate dehydrogenase were assessed prior to and at 4-week intervals after treatment. Results: Median survival of all patients was 7 months (95% CI: 6–9 months). Overall, 33/60 (55%) patients reported a decrease in pain after the first radionuclide treatment. This percentage was similar for patients treated with Re168-HEDP and Sr89. Mean duration of reported pain response was 75 days (± 68 days) for Sr89 and 61 days (± 56 days) for Re186-HEDP, which was not significantly different. A lower blood hemoglobin concentration was associated with a lower pain response rate. In a multivariate Cox regression analysis, pain response to radionuclide treatment predicted longer survival after treatment. Conclusions: Pain response was present in 55% of patients. Serum hemoglobin concentration prior to radionuclide treatment predicted pain response for both Re186-HEDP and Sr89. A reduction in pain upon radionuclide treatment was associated with a longer survival after treatment.
Mild Brain Ischemia Produces Bladder Hyperactivity without Brain Damage in Rats
Authors: Yotsuyanagi Satoshi , Narimoto Kazutaka , Namiki Mikio
Published: July 2006

Introduction: The influence of brain ischemia without cerebral infarction on voiding function is unknown. To investigate the effects of a reduction in cerebral blood flow on voiding function, the influence of chronic cerebral hypoperfusion (CH) on bladder activity was examined in rats. Materials and Methods: CH was induced in each of 11 female Sprague-Dawley rats by anastomosis between the right external jugular vein and the right common carotid artery with partial obstruction of the left common carotid artery. Twelve intact animals comprised a control group. Voided volume per micturition was assessed in a metabolic cage for 24 h on weeks 2, 4, and 8. Eight weeks after the operation, the rats were tested in a hippocampus-related learning paradigm, the Morris water maze. Bladder activity was monitored in 13 rats with continuous infusion cystometrography (CMG) at 2 weeks. After evaluation, the rats’ brains were stained by perfusion with 2% 2,3,5-triphenyltetrazolium chloride (TTC). Results: Voided volume per micturition was significantly reduced and voiding frequency was significantly increased in CH rats 2 weeks after CH as compared to the control group (p < 0.05). Bladder capacity on CMG of CH rats was significantly reduced 14 days after CH as compared to the controls (p < 0.05). Although TTC staining of the CH rat brain did not show cerebral infarction, CH induced impairment of water maze learning. Conclusions: These results indicate that mild forebrain ischemia without infarction results in the development of bladder hyperactivity and impairment of memory. Mild brain ischemia with aging may induce bladder overactivity in humans. Further studies of the nervous system related to bladder hyperactivity using this animal model may lead to pharmacological therapy or prevention of bladder overactivity in the aging individual with an unidentified origin of voiding dysfunction.
Recent Changes of the Incidence of Complete and Partial Mole in Chiba Prefecture
Authors: Matsui Hideo , Kihara Maki , Yamazawa Koji , Mitsuhashi Akira , Seki Katsuyoshi , Sekiya Souei
Published: July 2006

Aims: To study the changes of the incidence of complete mole (CM) and partial mole (PM) by 10-year age groups in Chiba Prefecture. Methods: All women registered as CM and PMs in Chiba Prefecture during these 18 years were included in this study. The diagnosis of CM and PM was based on the macroscopic and/or microscopic findings. The annual numbers of pregnancy were obtained from the Division of Statistics in Chiba Prefecture Government. Results: The incidence of CM at the upper and lower extremes of maternal age is higher than that of PM. The incidence of CM has decreased constantly at all maternal ages and significantly decreased in women of middle reproductive age (20–39 years old) since 1991, while that of PM has stayed constant during these 18 years. Conclusions: The incidence of CM and PM in Chiba Prefecture has become as low as that in Europe or the USA. These recent changes suggest that Japanese women may have lost the increased risk to ovulate a nuclear or inactive oocytes, or the differential diagnosis between CM and PM may be obscured with the macroscopic and/or microscopic findings.
Analysis of Single Nucleotide Polymorphism Sites in Exon 4 of the p53 Gene Using High-Performance Liquid Chromatography/Electrospray Ionization/Tandem Mass Spectrometry
Authors: Song Renfang , Zhang Wenbing , Chen Huayong , Ma Huimin , Dong Yulian , Sheng Guoying , Zhou Zhen , Fu Jiamo
Published: July 2006

Three groups of four oligonucleotides with special single nucleotide polymorphism (SNP) sites in exon 4 of the p53 gene were analyzed with ion-pair, reversed-phase, high-performance liquid chromatography/electrospray ionization/tandem mass spectrometry. The retention order of four oligonucleotides with SNPs was C<G<A<T, regardless of whether the polymorphisms were at the 3′ end, the 5′ end, or the middle of the oligonucleotides. The charge state of the molecular ion affects the tandem mass spectra of the oligonucleotides. SNPs at the 3′ end can easily be identified from the fragmentation pattern of the 2- charge state, but not from the 3- charge state, especially from the w1 fragment. The single base may be taken as the symbol of the 5′ end SNP site derived from [M-3H]3−, but not from the [M-3H]2− charge state. The oligonucleotides with SNPs in the middle were also determined from the [M-2H]2− precursor ion.
Effects of 3- and 5-Year Treatment With Risedronate on Bone Mineralization Density Distribution in Triple Biopsies of the Iliac Crest in Postmenopausal Women
Authors: Zoehrer Ruth , Roschger Paul , Paschalis Eleftherios P , Hofstaetter Jochen G , Durchschlag Erich , Fratzl Peter , Phipps Roger , Klaushofer Klaus
Published: July 2006

Long-term effects of risedronate on bone mineralization density distribution in triple transiliac crest biopsies of osteoporotic women were evaluated. In this double-blinded study, 3- and 5-year treatment with risedronate increased the degree and homogeneity of mineralization without producing hypermineralization. These changes at the material level of bone could contribute to risedronate's antifracture efficacy.

Introduction: Risedronate, a nitrogen-containing bisphosphonate, is widely used in the treatment of osteoporosis. It reduces bone turnover, increases BMD, and decreases fracture risk. To date, there are no data available on the long-term effects of risedronate on bone mineralization density distribution (BMDD) in humans.

Materials and Methods: Osteoporotic women enrolled in the VERT-NA trial received either risedronate (5 mg/day, orally) or placebo for up to 5 years. All subjects received calcium and vitamin D supplementation if deficient at baseline. Triple iliac crest biopsies were collected from a subset of these subjects at baseline and 3 and 5 years. BMDD was measured in these biopsies using quantitative backscattered electron imaging, and the data were also compared with a normal reference group.

Results: At baseline, both risedronate and placebo groups had a lower degree and a greater heterogeneity of mineralization as well as an increase in low mineralized bone compared with the normal reference group. The degree of mineralization increased significantly in the risedronate as well as in the placebo group after 3- and 5-year treatment compared with baseline. However, the degree of mineralization did not exceed that of normal. Three-year treatment with risedronate significantly increased the homogeneity of mineralization and slightly decreased low mineralized bone compared with placebo. Surprisingly with 5-year risedronate treatment, heterogeneity of mineralization increased compared with 3-year treatment, which might indicate an increase in newly formed bone.

Conclusions: Long-term treatment with risedronate affects the homogeneity and degree of mineralization without inducing hypermineralization of the bone matrix. These changes at the material level of the bone matrix may contribute to risedronate's antifracture efficacy in osteoporotic patients.
Strontium Ranelate Reduces the Risk of Vertebral and Nonvertebral Fractures in Women Eighty Years of Age and Older
Authors: Seeman Ego , Vellas Bruno , Benhamou Claude , Aquino Jean Pierre , Semler Jutta , Kaufman Jean Marc , Hoszowski Krzysztof , Varela Alfredo Roces , Fiore Carmelo , Brixen Kim , Reginster Jean Yves , Boonen Steven
Published: July 2006

Strontium ranelate produces an early and sustained reduction of both vertebral and nonvertebral fractures in patients ≥80 years of age.

Introduction: About 25–30% of the population burden of all fragility fractures in the community arise from women ≥80 years of age, because this population is at high risk for all types of fracture, particularly nonvertebral fractures. Despite this, evidence that therapies reduce the risk of both vertebral and nonvertebral fractures in this group is lacking. The aim of this study was to determine whether strontium ranelate, an agent that reduces the risk of vertebral and nonvertebral fractures in postmenopausal women >50 years of age, also reduces fractures in the elderly.

Materials and Methods: An analysis based on preplanned pooling of data from two international, phase III, randomized, placebo-controlled, double-blind studies (the Spinal Osteoporosis Therapeutic Intervention [SOTI] and TReatment Of Peripheral OSteoporosis [TROPOS]) included 1488 women between 80 and 100 years of age followed for 3 years. Yearly spinal X-rays were performed in 895 patients. Only radiographically confirmed nonvertebral fractures were included.

Results: Baseline characteristics did not differ in placebo and treatment arms. In the intent-to-treat analysis, the risk of vertebral, nonvertebral, and clinical (symptomatic vertebral and nonvertebral) fractures was reduced within 1 year by 59% (p = 0.002), 41% (p = 0.027), and 37% (p = 0.012), respectively. At the end of 3 years, vertebral, nonvertebral, and clinical fracture risks were reduced by 32% (p = 0.013), 31% (p = 0.011), and 22% (p = 0.040), respectively. The medication was well tolerated, and the safety profile was similar to that in younger patients.

Conclusions: Treatment with strontium ranelate safely reduces the risk of vertebral and nonvertebral fractures in women with osteoporosis ≥80 years of age. Even in the oldest old, it is not too late to reduce fracture risk.
A Novel Mutation (K378X) in the Sequestosome 1 Gene Associated With Increased NF-κB Signaling and Paget's Disease of Bone With a Severe Phenotype
Authors: Rea Sarah L , Walsh John P , Ward Lynley , Yip Kirk , Ward Bryan K , Kent G Neil , Steer James H , Xu Jiake , Ratajczak Thomas
Published: July 2006

Sequestosome 1/p62 (p62) mutations are associated with PDB; however, there are limited data regarding functional consequences. We report a novel mutation in exon 7 (K378X) in a patient with polyostotic Paget's disease of bone. p62 mutants increased NF-κB activation and significantly potentiated osteoclast formation and bone resorption in human primary cell cultures.

Introduction: Sequestosome 1/p62 (p62) mutations are associated with Paget's disease of bone (PDB); however, there are limited data regarding functional consequences. One report has linked the common P392L mutation in the p62 ubiquitin binding associated (UBA) domain with increases in NF-κB activity, a transcription factor essential for osteoclastogenesis. To further clarify the functional impact of p62 mutations associated with PDB, we assessed the effect of p62 mutation (a novel mutation: K378X, and previously reported mutations: P392L and E396X) on RANK-induced NF-κB activation and compared this with the effect of wildtype p62. In addition, we studied the effect of p62 mutation on osteoclast formation and bone resorption.

Materials and Methods: We performed co-transfection experiments with expression plasmids for p62 (wildtype or mutated) and RANK and an NF-κB luciferase reporter gene. Luciferase activities were recorded after addition of luciferin to cellular lysates. RAW264.7 cells stably expressing enhanced green fluorescent protein (EGFP)-tagged p62 (wildtype, K378X, or P392L) or EGFP alone were assessed for changes in cell proliferation. Additionally, these cells were stimulated with RANKL to produce osteoclast-like cells (OLCs). Primary human monocytes collected from the K378X-affected patient and a control subject were stimulated to form OLCs and bone resorption data were obtained.

Results: The novel mutation introduces a premature stop codon in place of Lys-378 and thereby eliminates the entire p62 UBA domain; this and two additional natural mutations (P392L, E396X) increased NF-κB activation compared with wildtype p62. Wildtype p62 consistently inhibited NF-κB activation compared with empty vector. UBA mutations (K378X and P392L) significantly increased the number of OLCs formed in response to RANKL and also the number of nuclei of the OLCs. K378X-affected human monocytes formed more OLCs with more nuclei and increased bone resorption compared with control monocytes.

Conclusions: Our data show that mutation of the p62 UBA domain results in increased activation of NF-κB and osteoclast formation and function compared with wildtype p62. These results may partially explain the mechanism by which p62 mutation contributes to the pathogenesis of PDB.
Fluid Shear Stress Induces Less Calcium Response in a Single Primary Osteocyte Than in a Single Osteoblast: Implication of Different Focal Adhesion Formation
Authors: Kamioka Hiroshi , Sugawara Yasuyo , Murshid Sakhr A , Ishihara Yoshihito , Honjo Tadashi , Takano-Yamamoto Teruko
Published: July 2006

The immediate calcium response to fluid shear stress was compared between osteocytes and osteoblasts on glass using real-time calcium imaging. The osteoblasts were responsive to fluid shear stress of up to 2.4 Pa, whereas the osteocytes were not. The difference in flow-induced calcium may be related to differences in focal adhesion formation.

Introduction: To explore the immediate response to mechanical stress in a bone cell population, we examined flow-induced calcium transients. In addition, the involvement of focal adhesion-related calcium transients in response to fluid flow in the cells was studied.

Materials and Methods: Bone cells were isolated from 16-day-old embryonic chicken calvaria by serial treatment with EDTA and collagenase. Single cells on glass without intercellular connections were subjected to fluid flow, and intracellular calcium concentration was measured using imaging with fluo-3. The identification of cell populations in the same field was performed with a chick osteocyte-specific antibody, OB7.3, and an alkaline phosphatase substrate, ELF-97, for osteoblast identification afterward. Immunofluorescence staining of vinculin was performed to visualize focal adhesions.

Results: The percentage of cells responding to fluid shear stress at 1.2 Pa was 5.5% in osteocytes, 32.4% in osteoblasts, and 45.6% in OB7.3/ELF-97-negative cells. Furthermore, osteoblasts and OB7.3/ELF-97-negative cells were more responsive to 2.4 Pa than 1.2 Pa, whereas osteocytes were less responsive. The elevation of calcium transients over baseline did not show any significant differences in the populations. To elucidate the mechanism accounting for the fact that single osteocytes are less sensitive to fluid shear stress of up to 2.4 Pa than osteoblasts, we studied focal adhesion-related calcium transients. First, we compared focal adhesion formation between osteocytes and osteoblasts and found a larger number of focal adhesions in osteoblasts than in osteocytes. Next, when the cells were pretreated with GRGDS (0.5 mM) before flow treatment, a significant reduction of calcium transients in osteoblasts (18%) was observed, whereas calcium transients in osteocytes were not changed by GRGDS. Control peptide GRGES did not reduce the calcium transients in either cell type. Furthermore, we confirmed that osteoblasts in calvaria showed a marked formation of vinculin plaques in the periphery of the cells. However, osteocytes in calvaria showed faint vinculin plaques only at the base of the processes.

Conclusions: On glass, single osteocytes are less sensitive to fluid shear stress up to 2.4 Pa than osteoblasts. The difference in calcium transients might be related to differences in focal adhesion formation. Shear stress of a higher magnitude or direct deformation may be responsible for the mechanical response of osteocytes in bone.
Disease Status in Autosomal Dominant Osteopetrosis Type 2 Is Determined by Osteoclastic Properties
Authors: Chu Kang , Snyder Richard , Econs Michael J
Published: July 2006

Asymptomatic gene carriers and clinically affected ADO2 subjects have the same ClCN7 mutation. We examined osteoclastic bone resorption in vitro as well as osteoclast formation, several markers, acid secretion, and cytoskeletal structure. We found that ADO2 expression results from osteoclast specific properties.

Introduction: Autosomal dominant osteopetrosis type II (ADO2) is a heritable osteosclerotic disorder that results from heterozygous mutations in the ClCN7 gene. However, of those individuals with a ClCN7 mutation, one third are asymptomatic gene carriers who have no clinical, biochemical, or radiological manifestations. Disease severity in the remaining two thirds is highly variable.

Materials and Methods: Human peripheral blood mononuclear cells were isolated and differentiated into osteoclasts by stimulation with hRANKL and human macrophage-colony stimulating factor (hM-CSF). Study subjects were clinically affected subjects, unaffected gene carriers, and normal controls (n = 6 in each group). Pit formation, TRACP staining, RANKL dose response, osteoclast markers, acid secretion, F-actin ring, and integrin αvβ3 expression and co-localization were studied.

Results: Osteoclasts from clinically affected subjects had severely attenuated bone resorption compared with those from normal controls. However, osteoclasts from unaffected gene carriers displayed similar bone resorption to those from normal controls. In addition, the resorption lacunae from both unaffected gene carriers and normal controls appeared much earlier and spread much more rapidly than those from clinically affected subjects. As time progressed, the distinction between clinically affected subjects and the other two groups increased. No significant difference was found in acidic secretion or osteoclast formation between the three groups. Osteoclast cytoskeletal organization showed no difference between the three groups but there was low cellular motility in clinically affected subjects.

Conclusions: Osteoclasts from the unaffected gene carriers, in contrast to those from the clinically affected subjects, functioned normally in cell culture. This finding supports the hypothesis that intrinsic osteoclast factors determine disease expression in ADO2. Further understanding of this mechanism is likely to lead to the development of new approaches to the treatment of clinically affected patients.
Bone Morphogenetic Proteins in Bone Stimulate Osteoclasts and Osteoblasts During Bone Development
Authors: Okamoto Mina , Murai Junko , Yoshikawa Hideki , Tsumaki Noriyuki
Published: July 2006

In this study, overexpression of noggin, a BMP antagonist, in developing bone caused significantly decreased osteoclast number as well as bone formation rate, resulting in increased bone mass with immature bone quality. BMP signaling plays important roles in normal bone development and regulation of bone resorption.

Introduction: Bone morphogenetic proteins (BMPs) act on various types of cells. Although involvement of BMP signals in osteoblast differentiation has been studied extensively, the effects of BMPs on osteoclasts have not been widely researched. Consequently, the net effects of BMPs on bone remain unclear. The purpose of this study was to delineate more fully the role of BMPs in skeletal biology.

Materials and Methods: We generated transgenic mice that express BMP4 or noggin in bone under the control of the 2.3-kb α1(I) collagen chain gene (Col1a1) promoter, and analyzed their bone phenotype. We also analyzed bone of transgenic mice expressing BMP4 specifically in cartilage.

Results: Mice overexpressing BMP4 in bone developed severe osteopenia with increased osteoclast number. Mice overexpressing noggin, a BMP antagonist, in bone showed increased bone volume associated with decreased bone formation rate and decreased osteoclast number. The noggin-transgenic tibias exhibited reduced periosteal bone formation and reduced resorption of immature bone in marrow spaces, associated with frequent fractures at the diaphysis. Co-culture of primary osteoblasts prepared from noggin-transgenic calvariae and wildtype spleen cells resulted in poor osteoclast formation, which was rescued by addition of recombinant BMP2, suggesting that noggin inhibits osteoclast formation by attenuating BMP activities in noggin-transgenic mice. The expression levels of Rankl were not decreased in primary osteoblasts from noggin transgenic mice. Immunoblot analysis showed increased phosphorylation of Smad1/5/8 in osteoclast precursor cells after 20-minute treatment with BMPs, suggesting that these cells are stimulated by BMPs. Mice overexpressing BMP4 in cartilage had enlarged bones containing thick trabeculae, possibly because of expansion of cartilage anlagen.

Conclusions: Overexpression of noggin in bone revealed that BMP signals regulate bone development through stimulation of osteoblasts and osteoclasts.
Heritability of Changes in Bone Size and Bone Mass With Age in Premenopausal White Sisters
Authors: Hui Siu L , Koller Daniel L , Foroud Tatiana M , Econs Michael J , Johnston C Conrad , Peacock Munro
Published: July 2006

Femoral neck area expands and BMD decreases in premenopausal women. We used longitudinal DXA measurements on 388 premenopausal white sisters to show significant heritability of the rates of change in femoral neck area, BMC, and BMD.

Introduction: Bone mass and structure are highly heritable. However, genetic effects on age-related changes in bone mass and structure in adults have been much less studied.

Materials and Methods: DXA measurements were made on 388 healthy white premenopausal sisters from 178 families. Rates of change in femoral neck area, BMC, and BMD, as well as body weight, were calculated from two measurements made an average of 5.7 years apart. Mixed models were used to test whether bone changes were related to age or weight change. Variance components models were used to estimate the heritability of the rates of change. A method was proposed to correct for the underestimation of heritabilities caused by measurement errors of the rates of change.

Results: Femoral neck area increased with age, whereas BMD decreased. All of the rates of change at the femoral neck were positively correlated with weight change, but the rates of femoral neck changes did not vary with age. Adjusted for weight change, change in femoral neck BMC became negative. Significant heritabilities (0.29–0.36) were detected for changes in femoral neck BMC, BMD, and area adjusted for weight changes. Correction for DXA measurement error in the rate estimates increased the heritability estimates (from 0.29–0.36 range to 0.37–0.64 range).

Conclusions: Rates of change are heritable for femoral neck area, BMC, and BMD in premenopausal white women.
Activation of Sirt1 Decreases Adipocyte Formation During Osteoblast Differentiation of Mesenchymal Stem Cells
Authors: Bäckesjö Carl-Magnus , Li Yan , Lindgren Urban , Haldosén Lars-Arne
Published: July 2006

In vitro, mesenchymal stem cells differentiate to osteoblasts when exposed to bone-inducing medium. However, adipocytes are also formed. We showed that activation of the nuclear protein deacetylase Sirt1 reduces adipocyte formation and promotes osteoblast differentiation.

Introduction: Mesenchymal stem cells (MSCs) can differentiate into osteoblasts, adipocytes, chondrocytes, and myoblasts. It has been suggested that a reciprocal relationship exists between the differentiation of MSCs into osteoblasts and adipocytes. Peroxisome proliferator-activated receptor γ2 (PPARγ2) is a key element for the differentiation into adipocytes. Activation of Sirt1 has recently been shown to decrease adipocyte development from preadipocytes through inhibition of PPARγ2.

Materials and Methods: We used the mouse mesenchymal cell line C3H10T1/2 and primary rat bone marrow cells cultured in osteoblast differentiation medium with or without reagents affecting Sirt1 activity. Adipocyte levels were analyzed by light microscopy and flow cytometry (FACS) after staining with Oil red O and Nile red, respectively. Osteoblast and adipocyte markers were studied with quantitative real-time PCR. Mineralization in cultures of primary rat bone marrow stromal cells was studied by von Kossa and alizarin red staining.

Results: We found that Sirt1 is expressed in the mesenchymal cell line C3H10T1/2. Treatment with the plant polyphenol resveratrol as well as isonicotinamide, both of which activate Sirt1, blocked adipocyte development and increased the expression of osteoblast markers. Nicotinamide, which inhibits Sirt1, increased adipocyte number and increased expression of adipocyte markers. Furthermore, activation of Sirt1 prevented the increase in adipocytes caused by the PPARγ-agonist troglitazone. Finally, activation of Sirt1 in rat primary bone marrow stromal cells increased expression of osteoblast markers and also mineralization.

Conclusions: In this study, we targeted Sirt1 to control adipocyte development during differentiation of MSCs into osteoblasts. The finding that resveratrol and isonicotinamide markedly inhibited adipocyte and promoted osteoblast differentiation may be relevant in the search for new treatment regimens of osteoporosis but also important for the evolving field of cell-based tissue engineering.
Instrumentation and early-age monitoring of concrete slabs
Authors: Austin S. A. , Robins P. J. , Bishop J. W.
Published: July 2006

This paper reports the instrumentation and monitoring techniques developed to improve understanding of the early-life behaviour of concrete industrial ground-floor slabs. Concrete strains, temperatures and joint movements were measured using vibrating-wire embedment strain gauges and thermistors, while ambient conditions were monitored to assess their effects. A reliable, accurate and easy-to-implement methodology, developed by instrumenting four types of floor slab (mesh-reinforced long strip, mesh-reinforced jointed large area pour, steel fibre-reinforced jointed large area pour and steel fibre-reinforced jointless large area pour) is described in detail, along with the further developments and modifications to the instrumentation. The methodology developed would be equally applicable to the early-life monitoring of other concrete structural elements. Some sample results of data gathered using the instrumentation methodology developed are included.
Design of solid reinforced concrete beams
Authors: Alnuaimi A. S. , Bhatt P.
Published: July 2006

Higher experimentally measured load resistance of a solid beam than a hollow one, with similar reinforcement, led to theoretical and experimental investigations on how the concrete core participates in stress resistance. Stress analysis was carried out on solid beams subjected to combined load of bending, shear and torsion. The beams were 300 × 300 mm in cross-section and 3800 mm in length. A three-dimensional in-house non-linear finite-element program was used for numerical analysis. From this analysis, a stress distribution was selected and used in computational experiments. It was found that the concrete core participates in the beams' behaviour and strength and cannot be ignored when combined loads of bending, shear and torsion are present. Its participation depends partly on the ratio of the torsion-to-bending moment and the ratio of shear stress caused by torsion to the shear stress caused by shear force. This computational study was followed by experimental testing of three beams designed using the selected stress distribution. Experimental results on the behaviour and ultimate load showed an acceptable agreement with the design values. Most of the longitudinal and transverse reinforcement in the front side, where shear stresses are additive, reached yield stress near failure load. The longitudinal reinforcement in the rear side, where shear stresses are subtractive, reached near yield stress but the transverse reinforcement did not reach yield stress, especially in the beams where bending was dominant. In addition, it was found that the smaller the ratio of torsion to bending, the larger the failure load.
A Disaggregate Approach to Aggregate Supply
Authors: Herdt Robert W.
Published: July 2006

Analyses of aggregate supply response in low‐income countries are practically nonexistent because of the index number problems and the difficulty of accounting for the effect of weather inherent in aggregate approaches. The disaggregate model presented here includes variables measuring weather, external capital inputs and technology, as well as crop and input prices, in individual crop supply functions. A technique for combining the direct and cross elasticities into a single aggregate elasticity is given, and the results obtained using the technique are outlined.
Specifying the Allocation of Income among Taxes, Consumption, and Savings in Linear Programming Models
Authors: Vandeputte J. M. , Baker C. B.
Published: July 2006

The proper specification of tax liabilities and consumption behavior is a significant problem in designing models for studying firm growth. A specification is suggested to represent allocation of income between consumption and savings in multiperiod linear programming models, while accounting for the effects of income tax. It is proposed as a method that conserves row space while providing flexibility in representing actual tax liabilities and actual consumption behavior in terms relevant for the study of growth alternatives and constraints.
Monthly Supply‐Demand Relationships for Fed Cattle and Hogs
Authors: Hayenga Marvin L. , Hacklander Duane
Published: July 2006

Monthly livestock supply, demand, and price variations reflect phenomena often not isolated in annual or quarterly time series analysis. A five‐equation model of cattle and hog monthly supply and demand behavior at the live market level was estimated via two‐stage least squares. Regular monthly demand fluctuations, income and population changes, and the slaughter supply per packer work day were found to affect cattle and hog prices. Marketing behavior by farmers was affected by feedlot inventories and the recent price change which apparently influenced their short‐run price expectations and subsequent marketing response.
Fish Cycles: A Harmonic Analysis
Authors: Waugh Frederick V. , Miller Morton M.
Published: July 2006

The landings and prices of many species of fish exhibit rather regular cyclical movements. The paper uses traditional methods of harmonic analysis to measure the lengths and amplitudes of landings and price cycles for cod, haddock, and blackback flounder landed at specific New England ports. Also discussed, briefly, is the periodicity of the total U. S. catch of all food fish. Findings demonstrate the occurrence of pronounced seasonal (12‐month) cycles, as well as cycles of several years' duration. The authors stress the value of cyclical analysis as a forecasting device and as an aid to understanding the nature of demand for fishery products.
A Model of Competitive Behavior in Food Industries
Authors: Handy C. R. , Padberg D. I.
Published: July 2006

While conventional industrial organization theory has been used to explain behavior of the food distribution oligopoly, a more careful analysis shows its behavior tendencies to be basically different from the manufacturing oligopoly. The larger distribution firm tends toward price competition while the smaller tends toward nonprice competition. The interaction between manufacturing and distribution oligopolies in the food industry provides a unique balance between progressiveness and economy which is not available in industries where the distribution oligopoly has not developed. This different consequence of large firms brings up some basic questions about antitrust objectives and philosophy.