Flux characteristics of cell culture medium in rectangular microchannels

Rectangular microchannels 50 μm high and 30, 40, 50, 60, or 70 μm wide were fabricated by adjusting the width of a gap cut in a polyethylene sheet 50 μm thick and sandwiching the sheet between an acrylic plate and a glass plate. Flux in the microchannels was measured under three different inner surface conditions: uncoated, albumin-coated, and confluent growth of rat fibroblasts on the bottom of the microchannels. The normalized flux in microchannels with cultured fibroblasts or albumin coating was significantly larger than that in the uncoated channels. The experimental data for all microchannels deviated from that predicted by classical hydrodynamic theory. At small aspect ratio the flux in the microchannels was larger than that predicted theoretically, whereas it became smaller at large aspect ratio. The aspect ratio rather than Reynolds number is the correct property for predicting the variation of the normalized friction factor. We postulate that two counteracting effects, rotation of large molecules and slip velocity at the corners of the microchannels, are responsible for the deviation. From these results we conclude that albumin coating should be carried out in the same way as when fabricating our integrating cell-culture system. The outcomes of this study are not only important for the design of our culture system, but also quite informative for general microfluidics.     

Towards microfabricated biohybrid artificial lung modules for chronic respiratory support

We have utilized soft lithography techniques to create three-dimensional arrays of blood microchannels and gas pathways in poly(dimethylsiloxane) (PDMS) that approach the microvascular scale of the natural lung. The blood microchannels were lined with endothelial cells in an effort to provide a non-thrombogenic surface that might ultimately reduce the need for systemic anticoagulation. A novel design and fabrication method were developed to create prototype modules for gas permeance and cell culture testing. The gas permeance modules contained 6 layers, four gas and two blood, while the modules for cell culture testing contained two layers of blood channels. The gas permeance of the modules was examined and maximum values of 9.16 × 10⁻⁶ and 3.55 × 10⁻⁵ mL/s/cm²/cmHg, for O₂ and CO₂ respectively, were obtained. Finally, endothelial cells were seeded and dynamically cultured in prototype cell culture modules. Confluent and viable cell monolayers were achieved after 10 days of perfusion.     

A blocking-free microfluidic fluorescence heterogeneous immunoassay for point-of-care diagnostics

In this article, a rapid, sensitive, and disposable microfluidic immunosensor is presented for point-of-care (POC) testing and clinical diagnosis. For the first time, the blocking process is eliminated from a microfluidic heterogeneous immunoassay by using protein A functionalized polydimethylsiloxane microchannels. The nonspecific binding of the assay is maintained around the chip background level by using a pair of antibodies with different affinity to protein A under optimized experimental conditions. C-reactive protein (CRP), a biomarker for inflammation and cardiovascular disease risk assessment, is selected as a model analyte to demonstrate the sensitivity of this blocking-free microfluidic heterogeneous immunoassay. A four parameter logistic function is used to model and assess the data. The limit of detection obtained is 0.54 μg/mL, which is lower than the cut-off value for clinical diagnosis. The overall assay is completed in 5 min. The protein A modified PDMS chips wet-stored at 4°C can maintain biofunctionality up to 14 months. The developed blocking-free microfluidic heterogeneous immunoassay will immediately provide benefits to most immunosensing microdevices targeted for POC diagnostics by shortening analysis time, simplifying fluid transportation, reducing sample consumption, and lowering waste generation.     

Assembly of bacteriophage P2 capsids from capsid protein fused to internal scaffolding protein

Most tailed bacteriophages with double-stranded DNA genomes code for a scaffolding protein, which is required for capsid assembly, but is removed during capsid maturation and DNA packaging. The gpO scaffolding protein of bacteriophage P2 also doubles as a maturation protease, while the scaffolding activity is confined to a 90 residue C-terminal “scaffolding” domain. Bacteriophage HK97 lacks a separate scaffolding protein; instead, an N-terminal “delta” domain in the capsid protein appears to serve an analogous role. We asked whether the C-terminal scaffolding domain of gpO could work as a delta domain when fused to the gpN capsid protein. Varying lengths of C-terminal sequences from gpO were fused to the N-terminus of gpN and expressed in E. coli. The presence of just the 41 C-terminal residues of gpO increased the fidelity of assembly and promoted the formation of closed shells, but the shells formed were predominantly small, 40 nm shells, compared to the normal, 55 nm P2 procapsid shells. Larger scaffolding domains fused to gpN caused the formation of shells of varying size and shape. The results suggest that while fusing the scaffolding protein to the capsid protein assists in shell closure, it also restricts the conformational variability of the capsid protein.     

A computational and experimental study inside microfluidic systems: the role of shear stress and flow recirculation in cell docking

In this paper, microfluidic devices containing microwells that enabled cell docking were investigated. We theoretically assessed the effect of geometry on recirculation areas and wall shear stress patterns within microwells and studied the relationship between the computational predictions and experimental cell docking. We used microchannels with 150 μm diameter microwells that had either 20 or 80 μm thickness. Flow within 80 μm deep microwells was subject to extensive recirculation areas and low shear stresses (<0.5 mPa) near the well base; whilst these were only presented within a 10 μm peripheral ring in 20 μm thick microwells. We also experimentally demonstrated that cell docking was significantly higher (p < 0.01) in 80 μm thick microwells as compared to 20 μm thick microwells. Finally, a computational tool which correlated physical and geometrical parameters of microwells with their fluid dynamic environment was developed and was also experimentally confirmed.     

Differential effect of storage on molecular and ultra-molecular dilutions of histamine

Objectives and design:  In this study, we examine the relationship between C5a and activation of cysteine aspartic acid protease 8 (caspase 8) in human umbilical vein endothelial cells (HUVEC). Materials or subjects:  Primary cultures of HUVEC were used. Treatments:  Recombinant human C5a (50 ng/ml) was used in the presence or absence of 10 μg/ml cycloheximide (CHX). Methods:  HUVEC were treated with C5a alone and in the presence of CHX, then monitored for cell viability, poly- ADP-ribose 1 (PARP-1) and caspase 8 activities. Gene and protein expressions were assessed for caspase 8 and the caspase 8 homologue, FLICE –inhibitory protein (cFLIP). Results:  We found a 43.1 ± 6.9 percent reduction in viability of HUVEC stimulated for 18 h with 50 ng/ml C5a in the presence of 10 μg/ml CHX (p < 0.05). In contrast, the cell viability of cells stimulated for 18 h with 50 ng/ml C5a or 10 μg/ml CHX alone was not significantly different compared to the non-stimulated control. Treatment of HUVEC with C5a induced an increase in caspase 8 activity but did not significantly affect cFLIP levels. Conclusions:  These data suggest caspase 8 activation induced by C5a leads to cell death if protein synthesis of antiapoptotic protein(s) is blocked. Received 23 July 2008; returned for revision 10 September 2008; received for final revision 29 September 2008; accepted by M. Parnham 18 September 2008     

17β-Estradiol-induced enhancement of estrogen receptor biosynthesis via MAPK pathway in mouse skeletal muscle myoblasts

The skeletal muscle is one of the important target tissues for the actions of estrogen via both nuclear and extranuclear (non-genomic) pathways. However, there is a paucity of information about the receptor (ER) involved. The aim of this study was thus to explore the ER expression in skeletal muscle, and the influence of estrogen on it, by using C2C12 myoblasts derived from mouse skeletal muscle. Significant expression of a ~66-kD protein immunoreactive to ER type α (ERα) monoclonal antibody, which was comparable to that in ovary, was detected in the whole-cell (total) and nucleus-free (nonnuclear) fractions of C2C12 myoblasts. The expression level of these ER proteins increased in several hours with treatment with 17β-estradiol (E2), which was preceded by the elevation of the ER mRNA level. This increase appeared to reflect the acceleration of de novo synthesis of ER protein, as proved by the ³⁵S-methionine immunoprecipitation method. A similar extent of fast increase in ER expression was also induced by a membrane-impermeable, BSA-conjugated estradiol (E2-BSA). Unexpectedly, the E2-induced increases in total and nonnuclear ER were further enhanced by the classic ER antagonists tamoxifen and ICI182,780 in a wide concentration range, implying some structural difference of the involved ER from the classical one. Treatment with the ERK1/2 inhibitor, PD98059 (10 μM), or the p38 MAPK-specific inhibitor, SB203580 (10 μM), greatly inhibited the E2-induced ER increase, while the protein kinase C (PKC) activator TPA (1 μM) enhanced it. These results collectively suggest that C2C12 skeletal myoblasts express a high level of ER, a considerable part of which is extranuclear. Further, the expression of ER in these cells may be significantly upregulated by estrogen itself via increased biosynthesis linked to membrane-bound ER and downstream MAPK-mediated signaling pathways. Electronic supplementary material  The online version of this article (doi:) contains supplementary material, which is available to authorized users.     

The PWC170: comparison of different stage lengths in 11–16 year olds

It is unknown how the estimation of aerobic fitness in children and adolescents compares among physical working capacity (PWC) protocols with different stage lengths. The purpose of this study was twofold: (1) compare PWC tests with 2-, 3-, and 6-min stage lengths in youth, and (2) examine the relationship between PWC at a heart rate (HR) of 170 beats min⁻¹ (PWC170) and peak oxygen consumption (VO₂peak). Fifty youth (31 m, 19 f), aged 11–16 years participated. Each participant visited the laboratory twice and performed three PWC tests (2-, 3-, 6-min stages) on a cycle ergometer. Tests usually consisted of three stages of increasing loads with the goal of reaching HR ≥ 165 beats min⁻¹. Individual regression lines were created to predict workload at HR = 170 beats∙min⁻¹ for each test. Participants completed two VO_2peak tests, both running and cycling. Repeated measures ANOVA was used to compare PWC170 values. Pearson correlation was used to assess the relationships between VO_2peak and power output for different PWC170 stage lengths. The three PWC170 protocols differ significantly; therefore, it is not advisable to directly compare results from different protocols. Furthermore, PWC170 showed moderate associations with VO_2peak, with the 2-min protocol showing the best correlation.     

Control of 3-dimensional collagen matrix polymerization for reproducible human mammary fibroblast cell culture in microfluidic devices

Interest in constructing a reliable 3-dimensional (3D) collagen culture platform in microfabricated systems is increasing as researchers strive to investigate reciprocal interaction between extracellular matrix (ECM) and cells under various conditions. However, in comparison to conventional 2-dimensional (2D) cell culture research, relatively little work has been reported about the polymerization of collagen type I matrix in microsystems. We, thus, present a study of 3D collagen polymerization to achieve reproducible 3D cell culture in microfluidic devices. Array-based microchannels are employed to efficiently examine various polymerization conditions, providing more replicates with less sample volume than conventional means. Collagen fibers assembled in microchannels were almost two-times thinner than those in conventional gels prepared under similar conditions, and the fiber thickness difference influenced viability and morphology of embedded human mammary fibroblast (HMF) cells. HMF cells contained more actin stress fibers and showed increased viability in 3D collagen matrix composed of thicker collagen fibers. Relatively low pH of the collagen solution within a physiological pH range (6.5–8.5) and pre-incubation at low temperature (～4 °C) before polymerization at 37 °C allow sufficient time for molecular assembly, generating thicker collagen fibers and enhancing HMF cell viability. The results provide the basis for improved process control and reproducibility of 3D collagen matrix culture in microchannels, allowing predictable modifications to provide optimum conditions for specific cell types. In addition, the presented method lays the foundation for high throughput 3D cellular screening.     

Temperature Pretreatment Alters the Polarization Response of Human Neutrophils to the Chemoattractant N-formyl-Met-Leu-Phe

Neutrophils present a polarized morphology upon stimulation of chemoattractants, which play a vital role in host-defense mechanisms. Many studies have been published on neutrophil polarization, in which three different temperatures pretreatment (4°C, 25°C and 37°C) have been used. However, no study has investigated whether different temperature pretreatments affect neutrophil polarization. In the current study, we examined the effects of 4°C, 25°C and 37°C pretreatment temperatures on short-term (1 or 3 min) chemoattractant-induced polarization. Human neutrophils were polarized upon the stimulation of N-formyl-Met-Leu-Phe (fMLP) after pretreated by different temperature. The morphological changes of the neutrophils were investigated under the microscopy. The F-actin polymerization was determined by immunological histological chemistry. There were more head–tail polarized cells (>50% of the cells) in the 25°C and 37°C pretreatment groups than in the 4°C group (32.4%). The average lengths of the pseudopod were 3.2 ± 1.1 μm (n = 17), 5.3 ± 2.1 μm (n = 12) and 7.4 ± 2.7 μm (n = 21) in the 4°C, 25°C and 37°C pretreatment groups, respectively; the 4°C and 37°C pretreatment groups were statistically different (P < 0.05). Additionally, there was a statistically significant difference in the pseudopod extension rate among the three groups, as well as the Lamellipod percentage between the 4°C group and the other two groups within 1 min of stimulation with fMLP. This study demonstrates that different temperature pretreatments affect neutrophil polarization upon short-term stimulation with fMLP. Dongliang Zhao, Xiaojing Meng, Chunqing Cai equally contributed to this paper.     

Immunochemical Composition of Cryoglobulins Generated in Stroke

Introduction  Cryoglobulins are abnormal immune complexes where both the antigens and the antibodies are immunoglobulins. The ability of cryoglobulins to bind C-reactive protein and low density lipoproteins, activate complement, and stimulate production of tumor necrosis factor-α generates interest in studying cryoglobulins in ischemic stroke. Materials and Methods  We determined blood levels of cryoglobulins in patients with ischemic stroke at different time points of stroke onset and identified the composition of cryoglobulins isolated from the blood on the first day of stroke onset. Results  On days 1–14, significantly elevated levels of cryoglobulins were detected with the maximum level on day 3. Discussion  Determination of immunoglobulin (Ig) content of cryoglobulins revealed the presence of a mixture of polyclonal IgG, IgA, and IgM, C3 complement protein and its activation split products, C1q complement protein, pathogenic lipoprotein-X, and β-lipoprotein. Conclusion  We suggest that cryoglobulins are involved in post-ischemic inflammatory response through activation of the complement cascade and cytokines production.     

Multichannel oscillatory-flow multiplex PCR microfluidics for high-throughput and fast detection of foodborne bacterial pathogens

In the field of continuous-flow PCR, the amplification throughput in a single reaction solution is low and the single-plex PCR is often used. In this work, we reported a flow-based multiplex PCR microfluidic system capable of performing high-throughput and fast DNA amplification for detection of foodborne bacterial pathogens. As a demonstration, the mixture of DNA targets associated with three different foodborne pathogens was included in a single PCR solution. Then, the solution flowed through microchannels incorporated onto three temperature zones in an oscillatory manner. The effect factors of this oscillatory-flow multiplex PCR thermocycling have been demonstrated, including effects of polymerase concentration, cycling times, number of cycles, and DNA template concentration. The experimental results have shown that the oscillatory-flow multiplex PCR, with a volume of only 5 μl, could be completed in about 13 min after 35 cycles (25 cycles) at 100 μl/min (70 μl/min), which is about one-sixth of the time required on the conventional machine (70 min). By using the presently designed DNA sample model, the minimum target concentration that could be detected at 30 μl/min was 9.8 × 10⁻² ng/μl (278-bp, S. enterica), 11.2 × 10⁻² ng/μl (168-bp, E. coli O157: H7), and 2.88 × 10⁻² ng/μl (106-bp, L. monocytogenes), which corresponds to approximately 3.72 × 10⁴ copies/μl, 3.58 × 10⁴ copies/μl, and 1.79 × 10⁴ copies/μl, respectively. This level of speed and sensitivity is comparable to that achievable in most other continuous-flow PCR systems. In addition, the four individual channels were used to achieve multi-target PCR analysis of three different DNA samples from different food sources in parallel, thereby achieving another level of multiplexing.     

Living at high altitude in combination with sea-level sprint training increases hematological parameters but does not improve performance in rats

The regimen of aerobic training at sea level with recovery at high altitude has been used by athletes to improve performance. However, little is known about the effects of hypoxia when combined with sprint interval training on performance. The aim of the present study was to determine the effect of a “living high-sprint training low” strategy on hemoglobin, hematocrit and erythropoietin levels in rats. We also wanted to test whether the addition of a hypoxic stress to the program of daily treadmill running at high speeds induces expressional adaptations in skeletal muscle and affects performance. The protein content of peroxisome proliferator-activated receptor-γ coactivator-1α (PGC-1α), cytochrome C, pyruvate dehydrogenase kinase (PDK1), heat shock protein 70 (HSP70), manganese superoxide dismutase (MnSOD) and citrate synthase activity were determined in different muscle fiber types in our animals (red and white gastrocnemius muscle). We also determined the maximal aerobic velocity (MAV) before and after the training period. A total of 24 male Wistar rats (3 month old) were randomly divided into four experimental groups: the normoxic control group (n = 6), the normoxic trained group (n = 6), the hypoxic control group (12 h pO₂ 12%/12 h pO₂ 21%) (n = 6) and the hypoxic trained group (12 h pO₂ 12%/12 h pO₂ 21%). Living in normobaric hypoxia condition for 21 days significantly increased hemoglobin, hematocrit and erythropoietin levels in both the rest and the trained groups. The trained animals (normoxia and hypoxia) significantly increased their maximal aerobic velocity. No changes were found in the skeletal muscle in PGC-1α, cytochrome C, PDK1, HSP70, MnSOD protein content and in the citrate synthase activity in any experimental group. Regardless of whether it is combined with sprint interval training or not, after 21 days of living at high altitude we found a significant increase in the hematological values determined in our study. However, contrary to our starting hypothesis, the combination of normobaric hypoxia and sprint training did not improve MAV in our animals.     

The in vitro and in vivo anti-metastatic efficacy of oxythiamine and the possible mechanisms of action

This study examined the anti-metastatic effects of oxythiamine (OT) both in cell culture and in vivo. Cell culture results revealed that OT (0–20 μM) significantly inhibited the invasion and migration (IC₅₀ = 8.75 μM) of Lewis lung carcinoma (LLC) cells. These effects of OT were accompanied by the inhibition of metalloproteinases-2 and -9 (MMP-2, MMP-9), urokinase-type plasminogen activator (uPA) activities and by the increases in protein expression of tissue inhibitors of metalloproteinases-1 and -2 (TIMP-1, TIMP-2). We then implanted (s.c.) C57BL/6 mice with LLC cells and supplemented the mice with a low- or a high-dose of OT (250 or 500 mg/kg BW) daily for 5 wk. During the 5-wk period, OT supplementation decreased plasma MMP-2 activity in a dose-dependent manner, and this effect was significant after 4 wk of tumor cell implantation. Tumor metastasis was found to confine to the lungs of mice injected with the tumor cells. High-OT supplementation strongly lowered the number and area of tumors and inhibited protein expression of MMP-2 and MMP-9 in the lungs. In addition, high-OT supplementation markedly decreased the extent of proliferating cell nuclear antigen (PCNA) staining in the lungs. By contrast, OT supplementation increased TIMP-1 and -2 protein expression in the lungs. These results demonstrate that OT supplementation attenuates tumor cell metastasis, possibly via inhibition of protein expression of MMPs, extent of PCNA staining and via increase of proteins expression of TIMPs.     

Polyvinylpyrrolidone (PVP) Mitigates the Damaging Effects of Intracellular Ice Formation in Adult Stem Cells

The objective of this work was to assess the effect of 10% (w/v) polyvinylpyrrolidone (PVP) on the pattern of intracellular ice formation (IIF) in human adipose tissue derived adult stem cells (ASCs) in the absence of serum and other cryoprotective agents (CPAs). The freezing experiments were carried out using a fluorescence microscope equipped with a Linkam™ cooling stage using two cooling protocols. Both the cooling protocols had a common cooling ramp: cells were cooled from 20 °C to −8 °C at 20 °C/min and then further cooled to −13 °C at 1 °C/min. At this point we employed either cooling protocol 1: the cells were cooled from −13 °C to −40 °C at a pre-determined cooling rate of 1, 5, 10, 20, or 40 °C/min and then thawed back to 20 °C at 20 °C/min; or cooling protocol 2: the cells were re-warmed from −13 °C to −5 °C at 20 °C/min and then re-cooled at a pre-determined rate of 1, 5, 10, 20, or 40 °C/min to −40 °C. Almost all (>95%) of the ASCs frozen in 1× PBS and protocol 1 exhibited IIF. However, almost none (<5%) of the ASCs frozen in 1× PBS and protocol 2 exhibited IIF. Similarly, almost all (>95%) of the ASCs frozen in 10% PVP in PBS and protocol 1 exhibited IIF. However, ~0, ~40, ~47, ~67, and ~100% of the ASCs exhibited IIF when frozen in 10% PVP in PBS and utilizing protocol 2 at a cooling rate of 1, 5, 10, 20, or 40 °C/min, respectively.     

The Dynamics of the Weighting and Topographical Characteristics of the Excitatory Zones of the Receptive Fields of Neurons in the Cat Visual Cortex

Acute experiments on 22 anesthetized and immobilized cats used a time slice method to study changes in the maps of 83 on and/or off receptive fields in 47 neurons in field 17 of the visual cortex. The latent period of the appearance of the receptive field averaged 88 ± 5 msec and its persistence time was 192 ± 12 msec. During generation of responses, the area and weighting on the one hand and the location of its discharge center on the other, changed in a wavelike fashion up to three times in all the neurons studied, the duration of each wave averaging 95 ± 4 msec. The discharge center of the receptive field moved in a wavelike manner in 99% of cases, moving towards and away from the center of the overall map with a period of 67.3 ± 3 msec. In 72.5% of neurons, movement of the discharge center occurred via different trajectories, which were ellipses. The functional significance of changes in the receptive fields of striate neurons in relation to the dynamics of their detector properties are discussed, as are the possible mechanisms of these rearrangements.     

SIRT1, AMP-activated protein kinase phosphorylation and downstream kinases in response to a single bout of sprint exercise: influence of glucose ingestion

This study was designed to examine potential in vivo mechanisms of AMP-activated protein kinase (AMPK) phosphorylation inhibition and its downstream signaling consequences during the recovery period after a single bout of sprint exercise. Sprint exercise induces Thr¹⁷²-AMPK phosphorylation and increased PGC-1α mRNA, by an unknown mechanism. Muscle biopsies were obtained in 15 young healthy men in response to a 30-s sprint exercise (Wingate test) randomly distributed into two groups: the fasting (n = 7, C) and the glucose group (n = 8, G), who ingested 75 g of glucose 1 h before exercising to inhibit AMPKα phosphorylation. Exercise elicited different patterns of Ser²²¹-ACCβ, Ser⁴⁷³-Akt and Thr⁶⁴²-AS160 phosphorylation, during the recovery period after glucose ingestion. Thirty minutes after the control sprint, Ser⁴⁸⁵-AMPKα1/Ser⁴⁹¹-AMPKα2 phosphorylation was reduced by 33% coinciding with increased Thr¹⁷²-AMPKα phosphorylation (both, P < 0.05). Glucose abolished the 30-min Thr¹⁷²-AMPKα phosphorylation. Ser²²¹-ACCβ phosphorylation was elevated immediately following and 30 min after exercise in C and G, implying a dissociation between Thr¹⁷²-AMPKα and Ser²²¹-ACCβ phosphorylation. Two hours after the sprint, PGC-1α protein expression remained unchanged while SIRT1 (its upstream deacetylase) was increased. Glucose ingestion abolished the SIRT1 response without any significant effect on PGC-1α protein expression. In conclusion, glucose ingestion prior to a sprint exercise profoundly affects Thr¹⁷²-AMPKα phosphorylation and its downstream signaling during the recovery period.     

Proteins of the Plasmodium falciparum two transmembrane Maurer’s cleft protein family, PfMC-2TM, and the 130?kDa Maurer’s cleft protein define different domains of the infected erythrocyte intramembranous network

Plasmodium falciparum Maurer’s clefts participate in the transport of macromolecules within the cytoplasm, including the transport of virulence proteins to the erythrocyte membrane surface. We identified a family of genes PfMC-2TM encoding transmembrane proteins located within the intramembranous network of the infected erythrocyte using monoclonal antibody SP1C1. The distribution of the PfMC-2TM protein family within domains of the network was investigated by colocalization and confocal microscopy studies using monoclonal antibody SP1C1 specific for PFMC-2TM and monoclonal antibody SP1A6 specific for the130 kDa Maurer’s cleft protein. Peptide-specific antibodies were prepared against six peptides from different domains of PfMC-2TM and used with the Mabs, as well as known antibodies specific to Maurer’s clefts proteins (ring-expressed protein and membrane-associated histidine-rich protein 1), the erythrocyte membrane protein 1 (PfEMP-1), and serine-rich antigen in colocalization studies. We show that PfMC-2TM is located in the Maurer’s clefts throughout the intracellular blood stage, and immunoelectron microscopy shows domains of PfMC-2TM localized in the parasitophorous vacuole and parasitophorous vacuole membrane. The distribution of the 130 kDa Maurer’s cleft protein changes from within the parasite to the clefts during intracellular development as the parasite matures from young trophozoite to segmented schizont.     

Post-exercise protein synthesis rates are only marginally higher in type I compared with type II muscle fibres following resistance-type exercise

We examined the effect of an acute bout of resistance exercise on fractional muscle protein synthesis rates in human type I and type II muscle fibres. After a standardised breakfast (31 ± 1 kJ kg⁻¹ body weight, consisting of 52 Energy% (En%) carbohydrate, 34 En% protein and 14 En% fat), 9 untrained men completed a lower-limb resistance exercise bout (8 sets of 10 repetitions leg press and leg extension at 70% 1RM). A primed, continuous infusion of l-[ring-¹³C₆]phenylalanine was combined with muscle biopsies collected from both legs immediately after exercise and after 6 h of post-exercise recovery. Single muscle fibres were dissected from freeze-dried biopsies and stained for ATPase activity with pre-incubation at a pH of 4.3. Type I and II fibres were separated under a light microscope and analysed for protein-bound l-[ring-¹³C₆]phenylalanine labelling. Baseline (post-exercise) l-[ring-¹³C₆]phenylalanine muscle tissue labelling, expressed as (∂¹³C/¹²C), averaged −32.09 ± 0.28, −32.53 ± 0.10 and −32.02 ± 0.16 in the type I and II muscle fibres and mixed muscle, respectively (P = 0.14). During post-exercise recovery, muscle protein synthesis rates were marginally (8 ± 2%) higher in the type I than type II muscle fibres, at 0.100 ± 0.005 versus 0.094 ± 0.005%/h, respectively (P < 0.05), whereby rates of mixed muscle protein were 0.091 ± 0.005%/h. Muscle protein synthesis rates following resistance-type exercise are only marginally higher in type I compared with type II muscle fibres.     

Induction and decay of short-term heat acclimation

The purpose of this work was to investigate adaptation and decay from short-term (5-day) heat acclimation (STHA). Ten moderately trained males (mean ± SD age 28 ± 7 years; body mass 74.6 ± 4.4 kg; [(V)\dot]_{\textO 2\textpeak} \dot{V}_{{{\text{O}}_{ 2{\text{peak}}} }} 4.26 ± 0.37 l min⁻¹) underwent heat acclimation (Acc) for 90-min on 5-days consecutively (T _a = 39.5°C, 60% RH), under controlled hyperthermia (rectal temperature 38.5°C). Participants completed a heat stress test (HST) 1 week before acclimation (Acc), then on the 2nd and 8th day (1 week) following Acc (T _a = 35°C, 60% RH). Seven participants completed HSTs 2 and 3 weeks after Acc. HST consisted of 90-min cycling at 40% peak power output before an incremental performance test. Rectal temperature at rest (37.1 ± 0.4°C) was not lowered by Acc (95% CI −0.3 to 0.2°C), after 90-min exercise (38.6 ± 0.5°C) it reduced 0.3°C (−0.5 to −0.1°C) and remained at this level 1 week later (−0.5 to −0.1°C), but not two (0.1°C −0.4 to 0.5°C; n = 7) or 3 weeks. Similarly, heart rate after 90-min exercise (146 ± 21 b min⁻¹) was reduced (−13: −6 to −20 b min⁻¹) and remained at this level after 1 week (−13: −6 to −20 b min⁻¹) but not two (−9: 6 to −23 b min⁻¹; n = 7) or 3 weeks. Performance (746 s) increased 106 s: 59 to 152 s after Acc and remained higher after one (76 s: 31 to 122) but not two (15 s: −88 to 142 s; n = 7) or 3 weeks. Therefore, STHA (5-day) induced adaptations permitting increased heat loss and this persisted 1 week but not 2 weeks following Acc.