Computer-supported collaborative learning in the medical workplace: Students' experiences on formative peer feedback of a critical appraisal of a topic paper.

BACKGROUND: Medical workplace learning consists largely of individual activities, since workplace settings do not lend themselves readily to group learning. An electronic Learning Management with System Computer-Supported Collaborative Learning (CSCL) could enable learners at different workplace locations to discuss personal clinical experiences at a distance to enhance learning. AIM: To explore whether CSCL-enabled structured asynchronous discussions on an authentic task has additional value for learning in the medical workplace. METHODS: Between January 2008 and June 2010, we conducted an exploratory evaluation study among senior medical students that were engaged in clinical electives. Students wrote a Critical Appraisal of a Topic paper about a clinical problem they had encountered and discussed it in discipline homogeneous subgroups on an asynchronous forum in a CSCL environment. A mixed method design was used to explore students' perceptions of the CSCL arrangement with respect to their preparation and participation, the design and knowledge gains. We analysed the messages recorded during the discussions to investigate which types of interactions occurred. RESULTS: Students perceived knowledge improvement of their papers. The discussions were mostly task-focused. The students considered an instruction session and a manual necessary to prepare for CSCL. A high amount of sent messages and a high activity in discussion seem to influence scores on perceptions: 'participation' and 'knowledge gain' positively. CONCLUSION: CSCL appears to offer a suitable environment for peers to provide formative feedback on a Critical Appraisal of a Topic paper during workplace learning. The CSCL environment enabled students to collaborate in asynchronous discussions, which positively influenced their learning.

Disturbed nuclear orientation and cellular migration in A-type lamin deficient cells.

The nuclear lamina and the cytoskeleton form an integrated structure that warrants proper mechanical functioning of cells. We have studied the correlation between structural alterations and migrational behaviour in fibroblasts with and without A-type lamins. We show that loss of A-type lamins causes loss of emerin and nesprin-3 from the nuclear envelope, concurring with a disturbance in the connection between the nucleus and the cytoskeleton in A-type lamin-deficient (lmna -/-) cells. In these cells functional migration assays during in vitro wound healing revealed a delayed reorientation of the nucleus and the microtubule-organizing center during migration, as well as a loss of nuclear oscillatory rotation. These observations in fibroblasts isolated from lmna knockout mice were confirmed in a 3T3 cell line with stable reduction of lmna expression due to RNAi approach. Our results indicate that A-type lamins play a key role in maintaining directional movement governed by the cytoskeleton, and that the loss of these karyoskeletal proteins has important consequences for functioning of the cell as a mechanical entity.

Role of nuclear lamina-cytoskeleton interactions in the maintenance of cellular strength.

The response of individual cells to cellular stress is vital for cellular functioning. A large network of physically interconnected cellular components, starting from the structural components of the cells' nucleus, via cytoskeleton filaments to adhesion molecules and the extracellular matrix, constitutes an integrated matrix that functions as a scaffold allowing the cell to cope with mechanical stress. Next to a role in mechanical properties, this network also has a mechanotransductional function in the response to mechanical stress. This signaling route does not only regulate a rapid reorganization of structural components such as actin filaments, but also stimulates for example gene activation via NFkappaB and other transcription factors. The importance of an intact mechano-signaling network is illustrated by the physiological consequences of several genetic defects of cellular network components e.g. actin, dystrophin, desmin and lamins. These give rise to an impaired response of the affected cells to mechanical stress and often result in dystrophy of the affected tissue. Recently, the importance of the cell nucleus in cellular strength has been established. Several new interconnecting proteins, such as the nesprins that link the nuclear lamina to the cytoskeleton, have been identified. Furthermore, the function of nuclear lamins in determining cellular strength and nuclear stability was illustrated in lamin-knock-out cells. Absence of the A-type lamins or mutations in these structural components of the nuclear lamina lead to an impaired cellular response to mechanical stress and disturbances in cytoskeletal organization. In addition, laminopathies show clinical phenotypes comparable to those seen for diseases resulting from genetic defects in cytoskeletal components, further indicating that lamins play a central role in maintaining the mechanical properties of the cell.

Heat shock proteins and cardiovascular pathophysiology.

In the eukaryotic cell an intrinsic mechanism is present providing the ability to defend itself against external stressors from various sources. This defense mechanism probably evolved from the presence of a group of chaperones, playing a crucial role in governing proper protein assembly, folding, and transport. Upregulation of the synthesis of a number of these proteins upon environmental stress establishes a unique defense system to maintain cellular protein homeostasis and to ensure survival of the cell. In the cardiovascular system this enhanced protein synthesis leads to a transient but powerful increase in tolerance to such endangering situations as ischemia, hypoxia, oxidative injury, and endotoxemia. These so-called heat shock proteins interfere with several physiological processes within several cell organelles and, for proper functioning, are translocated to different compartments following stress-induced synthesis. In this review we describe the physiological role of heat shock proteins and discuss their protective potential against various stress agents in the cardiovascular system.

HSP70-mediated acceleration of translational recovery after stress is independent of ribosomal RNA synthesis.

HSP70 is known to protect cells against stressful events. In the present study, the hypothesis was investigated that elevated HSP70 levels protect RNA polymerase I during stress, leading to decreased inhibition of ribosomal RNA (rRNA) synthesis and accelerated recovery of protein translation after stress. To this end, transcriptional and translational activity was studied in H9c2 cells during recovery after a severe heat treatment (SHT, 1 h 45 degrees C) in the presence of elevated HSP70 levels. The latter was achieved by heat pretreatment or by adenovirus-mediated hsp70 gene transfer. Rates of transcription and translation were determined by measuring cellular 3H-labelled uridine and leucine incorporation, respectively. The two types of pretreatment did not affect basal rates of transcription and translation, immediately before SHT. During SHT, both transcriptional and translational rates dropped to less than 10% of basal levels in pretreated as well as non-pretreated cells. Two and four h after SHT, both transcriptional and translational rates were significantly higher in HSP70-overexpressing cells compared to non-pretreated cells. However, immediately after SHT, transcription rates were similarly depressed in non-pretreated and pretreated cells, showing that increased levels of HSP70 did not protect RNA polymerase I activity during SHT. Thus, the HSP70-mediated acceleration of translational recovery is not preceded in time by an enhanced recovery of rRNA synthesis. Therefore, the HSP70-mediated early recovery of protein synthesis after heat stress is independent of rRNA synthesis.

Heat pretreatment differentially affects cardiac fatty acid accumulation during ischemia and postischemic reperfusion.

We investigated whether the cardioprotection induced by heat stress (HS) pretreatment is associated with mitigation of phospholipid degradation during the ischemic and/or postischemic period. The hearts, isolated from control rats and from heat-pretreated rats (42 degrees C for 15 min) either 30 min (HS0.5-h) or 24 h (HS24-h) earlier, were subjected to 45 min of no-flow ischemia, followed by 45 min of reperfusion. Unesterified arachidonic acid (AA) accumulation was taken as a measure for phospholipid degradation. Significantly improved postischemic ventricular functional recovery was only found in the HS24-h group. During ischemia, AA accumulated comparably in control and both HS groups. During reperfusion in control and HS0.5-h hearts, AA further accumulated (control hearts from 82 +/- 33 to 109 +/- 51 nmol/g dry wt, not significant; HS-0.5h hearts from 52 +/- 22 to 120 +/- 53 nmol/g dry wt; P < 0.05). In contrast, AA was lower at the end of the reperfusion phase in HS24-h hearts than at the end of the preceding ischemic period (74 +/- 18 vs. 46 +/- 23 nmol/g dry wt; P < 0.05). Thus accelerated reperfusion-induced degradation of phospholipids in control hearts is completely absent in HS24-h hearts. Furthermore, the lack of functional improvement in HS0.5-h hearts is also associated with a lack of beneficial effect on lipid homeostasis. Therefore, it is proposed that enhanced membrane stability during reperfusion is a key mediator in the heat-induced cardioprotection.

Heat stress pretreatment mitigates postischemic arachidonic acid accumulation in rat heart.

Heat stress pretreatment of the heart is known to protect this organ against an ischemic/reperfusion insult 24 h later. Degradation of membrane phospholipids resulting in tissue accumulation of polyunsaturated fatty acids, such as arachidonic acid, is thought to play an important role in the multifactorial process of ischemia/reperfusion-induced damage. The present study was conducted to test the hypothesis that heat stress mitigates the postischemic accumulation of arachidonic acid in myocardial tissue, as a sign of enhanced membrane phospholipid degradation. The experiments were performed on hearts isolated from rats either 24 h after total body heat treatment (42 degrees C for 15 min) or 24 h after sham treatment (control). Hearts were made ischemic for 45 min and reperfused for another 45 min. Heat pretreatment resulted in a significant improvement of postischemic hemodynamic performance of the isolated rat hearts. The release of creatine kinase was reduced from 30 +/- 14 (control group) to 17 +/- 5 units/g wet wt per 45 min (heat-pretreated group) (p < or = 0.05). Moreover, the tissue content of the inducible heat stress protein HSP70 was found to be increased 3-fold 24 h after heat treatment. Preischemic tissue levels of arachidonic acid did not differ between heat-pretreated and control hearts. The postischemic ventricular content of arachidonic acid was found to be significantly reduced in heat-pretreated hearts compared to sham-treated controls (6.6 +/- 3.3. vs. 17.8 +/- 12.0 nmol/g wet wt). The findings suggest that mitigation of membrane phospholipid degradation is a potential mechanism of heat stress-mediated protection against the deleterious effects of ischemia and reperfusion on cardiac cells.

alpha B-crystallin and hsp25 in neonatal cardiac cells--differences in cellular localization under stress conditions.

Two members of the small heat shock protein family, alpha B-crystallin and hsp25, occur at high levels in the mammalian heart. To try and understand any differences in functioning, we compared their properties in cultured rat neonatal cardiac myocytes. Both proteins are stress-inducible, but the level of hsp25 is only slightly increased in cultured cardiac myocytes subjected to hyperthermic stress, while alpha B-crystallin levels even remain unchanged. Phosphorylation of alpha B-crystallin and to a lesser extent also of hsp25 is induced after the heat shock. Directly after heat stress, alpha B-crystallin and hsp25 are partly found in detergent-insoluble fractions, representing cytoskeletal/nuclear structures. Additionally, we show by confocal laser scanning microscopy that alpha B-crystallin and hsp25 become associated with sarcomeric structures directly after the heat shock, indicating a cytoskeletal protective function. Four to six hours after the heat shock, both proteins reoccupy their original positions in the cytoplasm again. In contrast to alpha B-crystallin, hsp25 not only translocates to the cytoskeleton but also migrates to positions inside the nucleus. Despite the fact that both proteins are normally part of the same complex, their behavior in neonatal cardiac myocytes appears to be very different. The sarcomeric association of alpha B-crystallin occurs under milder conditions and persists for a longer period of time in comparison with hsp25. Our findings suggest that alpha B-crystallin and hsp25 are both involved in protection of the cytoskeleton during stress situations in the heart, although in different manners. In addition, hsp25 also plays a role inside the nucleus.

Heat stress protects aged hypertrophied and nonhypertrophied rat hearts against ischemic damage.

To explore the effects of heat stress (HS) in aged hypertrophied and nonhypertrophied rat hearts, postischemic recovery was investigated 15 mo after aortic constriction (AoB) or sham operation (Sham). Twenty-four hours after HS (42 degrees C; 15 min) or control treatment (normothermia), global ischemia was induced for 20 min in isolated AoB hearts and for 20 or 30 min in Sham hearts. After HS, postischemic recovery after 20-min ischemia in AoB hearts and 30-min ischemia in Sham hearts, respectively, was significantly better than in corresponding controls. In AoB hearts, cardiac output (CO), left ventricular developed pressure (LVDP), and the positive maximal first derivative of left ventricular pressure (+dP/dtmax) recovered to 33 +/- 26 (means +/- SD), 87 +/- 5, and 72 +/- 12%, respectively, after HS and to 5 +/- 8, 22 +/- 39, and 17 +/- 29% of preischemic values, respectively, in controls. Postischemic arrhythmias were significantly reduced in HS hypertrophied hearts, but creatine kinase (CK) loss was not reduced. In Sham hearts subjected to 30 min ischemia, CO, LVDP, and +dP/dtmax recovered to 20 +/- 20, 75 +/- 8, and 59 +/- 15%, respectively, after HS and to 3 +/- 8, 21 +/- 32, and 16 +/- 32% of preischemic values, respectively, in controls. Duration of arrhythmias and CK loss were not reduced in the heated hearts. When Sham hearts were subjected to only 20-min ischemia, functional recovery was not different in HS and control hearts, indicating that HS pretreatment extends the ischemic interval before irreversible injury occurs in the heart. In all HS Sham hearts, the myocardial 72-kDa HS protein (HSP 70) content was significantly increased. However, in HS AoB hearts, HSP 70 levels were not significantly different from the values in the control hearts. These results indicate that HS pretreatment induces cardioprotection in aged hypertrophied and nonhypertrophied rat hearts, which, however, cannot be unequivocally related to increased HSP 70 tissue contents.

Calcium homeostasis in cardiomyocytes isolated from heat-shocked rats.

The cellular mechanism of heat shock-mediated cardioprotection is still under debate. Because heat pretreatment negatively affects the normoxic left ventricular contractile performance in vitro when the extracellular Ca2+ concentration ([Ca2+]o) is relatively low (0.65-1.25 mM), the intracellular Ca2+ homeostasis was studied in more detail in cardiomyocytes isolated from adult rats 24 h after heat stress (42 degrees C for 15 min) or anesthesia (control). Sensitivity to Ca2+ overload was assessed by exposure to veratridine (quiescent cells) or to [Ca2+]o ranging from 0.125 to 20 mM in quiescent and paced cardiomyocytes. The fraction of irreversibly hypercontracted cells was not different between groups. The fura-2 fluorescence ratio (I340/I380), which was used as a measure for cytoplasmic Ca2+ concentration ([Ca2+]i) in quiescent cells after exposure to [Ca2+]o (0.5-10 mM), was also not different between groups. Myofilament Ca2+ sensitivity was assessed in paced (0.5 Hz) cells by simultaneous measurement of [Ca2+]i transients and cell shortening. At stepwise increases of [Ca2+]o from 1 to 10 mM, these parameters were comparable between groups. The diastolic cell length shortened progressively and equally in both groups after increasing [Ca2+]o. However, within 2 min of return from 10 to 1 mM [Ca2+]o, cells from heat-shocked rats retained the same length, whereas cells from control rats contracted further (P = 0.05). These data suggest that heat stress improves relaxation after challenge with high [Ca2+]o.

Inability of the heat-shocked heart to adjust its pre-ischemic and post-ischemic performance to variable loading conditions.

The aim of the present study was to investigate whether the pre-ischemic and post-ischemic hemodynamic function of the heat-shocked rat heart is affected by changes in afterload and extracellular calcium concentrations ([Ca2+]e). Experiments were performed on isolated, ejecting Lewis rat hearts 24h after in vivo heat shock (LewHS) or anesthesia alone (Lewc). In vitro hearts were subjected to 60 min normoxic perfusion, 45 min global ischemia, and 60 min of reperfusion. Pre-ischemic and post-ischemic left ventricular performance was evaluated at [Ca2+]e ranging between 0.65 and 3.0 mM at afterloads of 8.0 kPa and 16.0 kPa. At 8.0 kPa, pre-ischemic function was comparable in LewHS and Lewc at [Ca2+]e equal to or above 2.25 mM. At lower [Ca2+]e, i.e., 0.65 and 1.25 mM, cardiac output (CO) was significantly lower in LewHS than in Lewc hearts. At 16.0 kPa, significantly lower CO values were found in LewHS than Lewc hearts at all [Ca2+]e levels. During post-ischemic reperfusion under basal conditions (8.0 kPa; [Ca2+]e = 2.25 mM) a significantly better recovery was observed in LewHS than Lewc hearts, persisting at [Ca2+]e equal to 1.25 mM. However, either by lowering [Ca2+]e to 0.65 mM or increasing afterload to 16.0 kPa (at all [Ca2+]e), heat shock-associated improvement of post-ischemic performance disappeared. In conclusion, pre-ischemic left ventricular performance of the isolated heat-shocked heart is depressed when it performs at low [Ca2+]e or against a relatively high afterload. The heat shock-mediated improvement of post-ischemic function is only present at relatively low afterload levels in combination with normal extracellular calcium concentrations.

Time-related normalization of maximal coronary flow in isolated perfused hearts of rats with myocardial infarction.

BACKGROUND: In the present study, we investigated the time dependency and regional differences of the vascular adaptation of the myocardium after myocardial infarction (MI) in rats. METHODS AND RESULTS: MI was induced by total occlusion of the left anterior descending coronary artery. Time-dependent adaptation of the coronary vasculature was determined by histological staining of endothelial cells and measurement of basal and maximal coronary flow at days 0, 4, 7, 21, 35, and 90 after surgery in isolated retrogradely perfused hearts of sham-operated and infarcted rats. Cardiac function was determined during anterograde perfusion. In a separate group of experiments, regional myocardial flow was measured with radiolabeled microspheres in sham-operated and infarcted hearts to determine local differences in adaptation. Basal coronary flow was completely normalized within 7 days, whereas maximal coronary flow was not normalized until 35 days after MI. Normal growth, as observed in sham-operated hearts, resulted in a parallel increase in coronary flow and tissue mass from day 7 to 35 after surgery. In contrast, the increase in coronary flow was lower than the hypertrophic response in the right ventricles and septa of infarcted hearts, whereas a parallel increase in tissue mass and coronary flow was observed in the left ventricles of these hearts. These functional data were supported by structural data that showed the presence of numerous and dilated vessels, especially in the border zone of the infarcted and noninfarcted tissue. CONCLUSIONS: These observations demonstrate that vessel growth, predominantly in the region adjacent to the infarcted zone, results in complete normalization of coronary vasodilatory capacity within 35 days after MI.

Mechanoperception and mechanotransduction in cardiac adaptation: mechanical and molecular aspects.

Cardiomyocytes grow in hypertrophy due to a net increase in the synthesis of proteins, especially contractile proteins, in the cell. There is abundant information about the molecular and biochemical changes involved in this process, but it is not completely understood how cells sense mechanical stimuli and how these stimuli are transferred into a biochemical signal inducing the growth response. This mechanotransduction most likely takes place at the cellular membrane. The resulting signal is transferred to the nucleus, where it can initiate alterations in gene expression.

A model approach to the adaptation of cardiac structure by mechanical feedback in the environment of the cell.

The uniformity of the mechanical load of the cardiac fibers in the wall is maintained by continuous remodeling. In this proposed model the myocyte changes direction in optimizing systolic sarcomere shortening. Early systolic stretch and contractility increases the mass of contractile proteins. Cyclic strain of the myocardial tissue diminishes passive stiffness, resulting in the control of ventricular end-diastolic volume. Utilizing these rules of remodeling in our mathematical model yields that the natural helical pathways of the myocardial fibers in the wall are formed automatically.

Heat shock improves ischemic tolerance of hypertrophied rat hearts.

The postischemic recovery of hypertrophied hearts was studied 24 h after total body hyperthermia. To this end, anesthetized aortic-banded and sham-operated rats were subjected to heat shock (AoBHS and ShamHS, respectively). Cardiac hypertrophy was induced 8 wk earlier. In isolated ejecting hearts, functional recovery after 45 min of global ischemia was poor and moderate in nonheated (control) hypertrophied (AoBC) and nonheated (control) nonhypertrophied (ShamC) hearts, respectively. Heat shock significantly improved postischemic recovery in both AoBHS and ShamHS hearts. This improvement of functional recovery was associated with a significant reduction of the duration of arrhythmias. In addition, coronary flow was significantly higher in both types of heat-shocked hearts than in the corresponding control hearts during the preischemic as well as the postischemic period. Postischemic endocardial flow, assessed using radioactive microspheres, was significantly improved in AoBHS hearts. Compared with the corresponding control hearts, the native endogenous catalase activity was not changed in AoBHS hearts but was significantly increased in ShamHS hearts. The present findings suggest that the postischemic functional improvement after total body hyperthermia can be explained by increased and more homogeneous myocardial perfusion, which may also reduce the duration of postischemic arrhythmias. This effect is especially beneficial for the hypertrophied heart, which is known to be extremely vulnerable to the ischemic insult probably caused by subendocardial underperfusion.