Effect of 28 days of creatine ingestion on muscle metabolism and performance of a simulated cycling road race

<p>Abstract</p> <p>Purpose</p> <p>The effects of creatine supplementation on muscle metabolism and exercise performance during a simulated endurance road race was investigated.</p> <p>Methods</p> <p>Twelve adult male (27.3 ± 1.0 yr, 178.6 ± 1.4 cm, 78.0 ± 2.5 kg, 8.9 ± 1.1 %fat) endurance-trained (53.3 ± 2.0 ml* kg^-1* min^-1, cycling ~160 km/wk) cyclists completed a simulated road race on a cycle ergometer (Lode), consisting of a two-hour cycling bout at 60% of peak aerobic capacity (VO_2peak) with three 10-second sprints performed at 110% VO_{2 peak}every 15 minutes. Cyclists completed the 2-hr cycling bout before and after dietary creatine monohydrate or placebo supplementation (3 g/day for 28 days). Muscle biopsies were taken at rest and five minutes before the end of the two-hour ride.</p> <p>Results</p> <p>There was a 24.5 ± 10.0% increase in resting muscle total creatine and 38.4 ± 23.9% increase in muscle creatine phosphate in the creatine group (<it>P </it>< 0.05). Plasma glucose, blood lactate, and respiratory exchange ratio during the 2-hour ride, as well as VO_{2 peak}, were not affected by creatine supplementation. Submaximal oxygen consumption near the end of the two-hour ride was decreased by approximately 10% by creatine supplementation (P < 0.05). Changes in plasma volume from pre- to post-supplementation were significantly greater in the creatine group (⁺14.0 ± 6.3%) than the placebo group (^-10.4 ± 4.4%; <it>P </it>< 0.05) at 90 minutes of exercise. The time of the final sprint to exhaustion at the end of the 2-hour cycling bout was not affected by creatine supplementation (creatine pre, 64.4 ± 13.5s; creatine post, 88.8 ± 24.6s; placebo pre, 69.0 ± 24.8s; placebo post 92.8 ± 31.2s: creatine vs. placebo not significant). Power output for the final sprint was increased by ~33% in both groups (creatine vs. placebo not significant).</p> <p>Conclusions</p> <p>It can be concluded that although creatine supplementation may increase resting muscle total creatine, muscle creatine phosphate, and plasma volume, and may lead to a reduction in oxygen consumption during submaximal exercise, creatine supplementation does not improve sprint performance at the end of endurance cycling exercise.</p

Analytic philosophy for biomedical research: the imperative of applying yesterday's timeless messages to today's impasses

The mantra that "the best way to predict the future is to invent it" (attributed to the computer scientist Alan Kay) exemplifies some of the expectations from the technical and innovative sides of biomedical research at present. However, for technical advancements to make real impacts both on patient health and genuine scientific understanding, quite a number of lingering challenges facing the entire spectrum from protein biology all the way to randomized controlled trials should start to be overcome. The proposal in this chapter is that philosophy is essential in this process. By reviewing select examples from the history of science and philosophy, disciplines which were indistinguishable until the mid-nineteenth century, I argue that progress toward the many impasses in biomedicine can be achieved by emphasizing theoretical work (in the true sense of the word 'theory') as a vital foundation for experimental biology. Furthermore, a philosophical biology program that could provide a framework for theoretical investigations is outlined

The HDAC Inhibitor FK228 Enhances Adenoviral Transgene Expression by a Transduction-Independent Mechanism but Does Not Increase Adenovirus Replication

The histone deacetylase inhibitor FK228 has previously been shown to enhance adenoviral transgene expression when cells are pre-incubated with the drug. Upregulation of the coxsackie adenovirus receptor (CAR), leading to increased viral transduction, has been proposed as the main mechanism. In the present study, we found that the highest increase in transgene expression was achieved when non-toxic concentrations of FK228 were added immediately after transduction, demonstrating that the main effect by which FK228 enhances transgene expression is transduction-independent. FK228 had positive effects both on Ad5 and Ad5/f35 vectors with a variety of transgenes and promoters, indicating that FK228 works mainly by increasing transgene expression at the transcriptional level. In some cases, the effects were dramatic, as demonstrated by an increase in CD40L expression by FK228 from 0.3% to 62% when the murine prostate cancer cell line TRAMP-C2 was transduced with Ad[CD40L]. One unexpected finding was that FK228 decreased the transgene expression of an adenoviral vector with the prostate cell-specific PPT promoter in the human prostate adenocarcinoma cell lines LNCaP and PC-346C. This is probably a consequence of alteration of the adenocarcinoma cell lines towards a neuroendocrine differentiation after FK228 treatment. The observations in this study indicate that FK228 enhances adenoviral therapy by a transduction-independent mechanism. Furthermore, since histone deacetylase inhibitors may affect the differentiation of cells, it is important to keep in mind that the activity and specificity of tissue- and tumor-specific promoters may also be affected

Bordetella pertussis Autotransporter Vag8 Binds Human C1 Esterase Inhibitor and Confers Serum Resistance

Bordetella pertussis employs numerous strategies to evade the immune system, including the ability to resist killing via complement. Previously we have shown that B. pertussis binds a complement regulatory protein, C1 esterase inhibitor (C1inh) to its surface in a Bvg-regulated manner (i.e. during its virulence phase), but the B. pertussis factor was not identified. Here we set out to identify the B. pertussis C1inh-binding factor. Using a serum overlay assay, we found that this factor migrates at approximately 100 kDa on an SDS-PAGE gel. To identify this factor, we isolated proteins of approximately 100 kDa from wild type strain BP338 and from BP347, an isogenic Bvg mutant that does not bind C1inh. Using mass spectrometry and bioinformatics, we identified the autotransporter protein Vag8 as the putative C1inh binding protein. To prove that Vag8 binds C1inh, vag8 was disrupted in two different B. pertussis strains, namely BP338 and 18–323, and the mutants were tested for their ability to bind C1inh in a surface-binding assay. Neither mutant strain was capable of binding C1inh, whereas a complemented strain successfully bound C1inh. In addition, the passenger domain of Vag8 was expressed and purified as a histidine-tagged fusion protein and tested for C1inh-binding in an ELISA assay. Whereas the purified Vag8 passenger bound C1inh, the passenger domain of BrkA (a related autotransporter protein) failed to do so. Finally, serum assays were conducted to compare wild type and vag8 mutants. We determined that vag8 mutants from both strains were more susceptible to killing compared to their isogenic wild type counterparts. In conclusion, we have discovered a novel role for the previously uncharacterized protein Vag8 in the immune evasion of B. pertussis. Vag8 binds C1inh to the surface of the bacterium and confers serum resistance

Internal mammary lymph node recurrence: rare but characteristic metastasis site in breast cancer

<p>Abstract</p> <p>Background</p> <p>To assess the frequency of IMLN recurrence, its associated risk factors with disease-free interval (DFI) and its predicting factors on overall survival time.</p> <p>Methods</p> <p>133 cases of breast cancer IMLN recurrence were identified via the computerized CT reporting system between February 2003 and June 2008, during which chest CT for patients with breast cancer (n = 8867) were performed consecutively at Cancer Hospital, Fudan University, Shanghai, China. Patients' charts were retrieved and patients' characteristics, disease characteristics, and treatments after recurrence were collected for analysis. The frequency was 1.5% (133/8867).</p> <p>Results</p> <p>IMLN recurrence was presented as the first metastatic site in 121 (91%) patients while 88 (66.2%) had other concurrent metastases. Typical chest CT images included swelling of the IMLN at the ipsilateral side with local lump and sternal erosion located mostly between the second and third intercostal space. The median disease-free interval (DFI) of IMLN recurrence was 38 months. The independent factors that could delay the IMLN recurrence were small tumor size (HR 0.5 95%CI: 0.4 - 0.8; <it>p </it>= 0.002), and positive ER/PR disease (HR 0.6, 95% CI: 0.4 - 0.9; <it>p </it>= 0.006). The median survival time after IMLN recurrence was 42 months, with a 5-year survival rate of 30%. Univariate analysis showed four variables significantly influenced the survival time: DFI of IMLN recurrence (p = 0.001), no concurrent distant metastasis (p = 0.024), endocrine therapy for patients with positive ER/PR (p = 0.000), radiotherapy (p = 0.040). The independent factors that reduced the death risk were no concurrent distant metastases (HR: 0.7, 95% CI: 0.4 - 0.9; <it>p </it>= 0.031), endocrine therapy for patients with positive ER/PR status (HR: 0.2, 95% CI: 0.1 - 0.5; <it>p </it>= 0.001) and palliative radiotherapy (HR: 0.3, 95% CI: 0.1- 0.9; <it>p </it>= 0.026).</p> <p>Conclusions</p> <p>The risk of IMLN recurrence is low and there are certain characteristics features on CT images. ER/PR status is both a risk factor for DFI of IMLN recurrence and a prognostic factor for overall survival after IMLN recurrence. Patients with only IMLN recurrence and/or local lesion have a good prognosis.</p

Rhabdomyolysis in Community Acquired Bacterial Sepsis – A Retrospective Cohort Study

BACKGROUND AND OBJECTIVES:Rhabdomyolysis is often associated with sepsis and gram positive bacterial pathogens are reported to be the most frequent cause of sepsis induced rhabdomyolysis. We report the pattern of infecting bacterial pathogens and associated causal factors in a South-Indian cohort. DESIGN, SETTING, PARTICIPANTS #ENTITYSTARTX00026; MEASUREMENTS:Retrospective cohort study of adult patients with community acquired bacterial sepsis complicated by rhabdomyolysis from March 2003--August 2008. Rhabdomyolysis was defined as serum creatine kinase >2000 IU/L. The study population was divided into group-I (sepsis with gram positive pathogens), group-II (sepsis with gram negative pathogens) and group-III (culture negative sepsis). RESULTS:103 patients (group I -15, group II- 34 and group III- 54) formed the study cohort. Mean age was 55 years and two-third had diabetes. Mean creatine kinase was 7114 IU/L and mean serum creatinine on admission was 2.4 mg/dl. Causative pathogen of sepsis was identified in 47.5%. Gram negative pathogens were more frequently (33%) associated with rhabdomyolysis than gram positive pathogens (14.5%). Lung was the commonest foci of sepsis (38.8%). 78.6% of the study population had one or more additional causal factor for rhabdomyolysis like statin intake, chronic alcoholism, hypokalemia, hypernatremia and hypophosphatemia. Mortality was 59%. CONCLUSIONS:Gram negative bacterial pathogens were more frequently associated with rhabdomyolysis than gram positive pathogens. Rhabdomyolysis in patients with sepsis is multifactorial and is associated with high mortality

Drosophila DNA polymerase theta utilizes both helicase-like and polymerase domains during microhomology-mediated end joining and interstrand crosslink repair

Double strand breaks (DSBs) and interstrand crosslinks (ICLs) are toxic DNA lesions that can be repaired through multiple pathways, some of which involve shared proteins. One of these proteins, DNA Polymerase theta (Pol theta), coordinates a mutagenic DSB repair pathway named microhomology-mediated end joining (MMEJ) and is also a critical component for bypass or repair of ICLs in several organisms. Pol theta contains both polymerase and helicase-like domains that are tethered by an unstructured central region. While the role of the polymerase domain in promoting MMEJ has been studied extensively both in vitro and in vivo, a function for the helicase-like domain, which possesses DNA-dependent ATPase activity, remains unclear. Here, we utilize genetic and biochemical analyses to examine the roles of the helicase-like and polymerase domains of Drosophila Pol theta. We demonstrate an absolute requirement for both polymerase and ATPase activities during ICL repair in vivo. However, similar to mammalian systems, polymerase activity, but not ATPase activity, is required for ionizing radiation-induced DSB repair. Using a site-specific break repair assay, we show that overall end-joining efficiency is not affected in ATPase-dead mutants, but there is a significant decrease in templated insertion events. In vitro, Pol theta can efficiently bypass a model unhooked nitrogen mustard crosslink and promote DNA synthesis following microhomology annealing, although ATPase activity is not required for these functions. Together, our data illustrate the functional importance of the helicase-like domain of Pol theta and suggest that its tethering to the polymerase domain is important for its multiple functions in DNA repair and damage tolerance

Tamoxifen and raloxifene modulate gap junction coupling during early phases of retinoic acid-dependent neuronal differentiation of NTera2/D1 cells

Gap junctions (GJ) represent a cellular communication system known to influence neuronal differentiation and survival. To assess a putative role of this system for neural effects of tamoxifen (TAM) and raloxifene (RAL), we used the human teratocarcinoma cell line NTera2/D1, retinoic acid (RA)-dependent neuronal differentiation of which is regulated by gap junctions formed of connexin43 (Cx43). As demonstrated by Western blot analysis, concentrations above 1 µmol/l for TAM, and 0.1 µmol/l for RAL lead to a temporary time- and concentration-dependent increase in Cx43 immunoreactivity, which reached a peak for TAM after 1 day and for RAL after 2 days. Immunocytochemical stainings revealed the increase in Cx43 immunoreactivity to result from an accumulation in intracellular compartments such as the Golgi apparatus or lysosomes. In addition, TAM and RAL were able to prevent the RA-dependent decrease of Cx43 immunoreactivity in NTera2/D1 cells, normally observed during neuronal differentiation. This suggested a suppression of neuronal differentiation to result from these substances. According to this, treatment of NTera2/D1 cells with 10 µmol/l TAM or RAL during weeks 1 and 2 of a 6 weeks RA-driven differentiation schedule impaired, whereas treatment during weeks 5 and 6 did not impair, neuronal differentiation of these cells. Modulation of GJ coupling between NTera2/D1 cells by TAM and RAL seems therefore to perturb early neuronal differentiation, whereas differentiated neurons in the mature brain seem to be not affected. These effects could be of importance for actions of TAM and RAL on early embryonic steps of nervous system formation

Structural and Functional Profiling of the Human Histone Methyltransferase SMYD3

The SET and MYND Domain (SMYD) proteins comprise a unique family of multi-domain SET histone methyltransferases that are implicated in human cancer progression. Here we report an analysis of the crystal structure of the full length human SMYD3 in a complex with an analog of the S-adenosyl methionine (SAM) methyl donor cofactor. The structure revealed an overall compact architecture in which the “split-SET” domain adopts a canonical SET domain fold and closely assembles with a Zn-binding MYND domain and a C-terminal superhelical 9 α-helical bundle similar to that observed for the mouse SMYD1 structure. Together, these structurally interlocked domains impose a highly confined binding pocket for histone substrates, suggesting a regulated mechanism for its enzymatic activity. Our mutational and biochemical analyses confirm regulatory roles of the unique structural elements both inside and outside the core SET domain and establish a previously undetected preference for trimethylation of H4K20