The Rac activator Tiam1 controls tight junction biogenesis in keratinocytes through binding to and activation of the Par polarity complex

The GTPases Rac and Cdc42 play a pivotal role in the establishment of cell polarity by stimulating biogenesis of tight junctions (TJs). In this study, we show that the Rac-specific guanine nucleotide exchange factor Tiam1 (T-lymphoma invasion and metastasis) controls the cell polarity of epidermal keratinocytes. Similar to wild-type (WT) keratinocytes, Tiam1-deficient cells establish primordial E-cadherin–based adhesions, but subsequent junction maturation and membrane sealing are severely impaired. Tiam1 and V12Rac1 can rescue the TJ maturation defect in Tiam1-deficient cells, indicating that this defect is the result of impaired Tiam1–Rac signaling. Tiam1 interacts with Par3 and aPKCζ, which are two components of the conserved Par3–Par6–aPKC polarity complex, and triggers biogenesis of the TJ through the activation of Rac and aPKCζ, which is independent of Cdc42. Rac is activated upon the formation of primordial adhesions (PAs) in WT but not in Tiam1-deficient cells. Our data indicate that Tiam1-mediated activation of Rac in PAs controls TJ biogenesis and polarity in epithelial cells by association with and activation of the Par3–Par6–aPKC polarity complex

The effect of asymmetric movement support on muscle activity during Lokomat guided gait in able-bodied individuals

BACKGROUND: To accommodate training for unilaterally affected patients (e.g. stroke), the Lokomat (a popular robotic exoskeleton-based gait trainer) provides the possibility to set the amount of movement guidance for each leg independently. Given the interlimb couplings, such asymmetrical settings may result in complex effects, in which ipsilateral activity co-depends on the amount of guidance offered to the contralateral leg. To test this idea, the effect of asymmetrical guidance on muscle activity was explored. METHODS: 15 healthy participants walked in the Lokomat at two speeds (1 and 2 km/h) and guidance levels (30% and 100%), during symmetrical (both legs receiving 30% or 100% guidance) and asymmetrical conditions (one leg receiving 30% and the other 100% guidance) resulting in eight unique conditions. Activity of the right leg was recorded from Erector Spinae, Gluteus Medius, Biceps Femoris, Semitendinosus, Vastus Medialis, Rectus Femoris, Medial Gastrocnemius and Tibialis Anterior. Statistical Parametric Mapping was used to assess whether ipsilateral muscle activity depended on guidance settings for the contralateral leg. RESULTS: Muscle output amplitude not only depended on ipsilateral guidance settings, but also on the amount of guidance provided to the contralateral leg. More specifically, when the contralateral leg received less guidance, ipsilateral activity of Gluteus Medius and Medial Gastrocnemius increased during stance. Conversely, when the contralateral leg received more guidance, ipsilateral muscle activity for these muscles decreased. These effects were specifically observed at 1 km/h, but not at 2 km/h. CONCLUSIONS: This is the first study of asymmetrical guidance on muscle activity in the Lokomat, which shows that ipsilateral activity co-depends on the amount of contralateral guidance. In therapy, these properties may be exploited e.g. to promote active contributions by the more affected leg. Therefore, the present results urge further research on the use of asymmetrical guidance in patient groups targeted by Lokomat training

Lokomat guided gait in hemiparetic stroke patients:the effects of training parameters on muscle activity and temporal symmetry

Purpose: The Lokomat is a commercially available robotic gait trainer, applied for gait rehabilitation in post-stroke hemiparetic patients. Selective and well-dosed clinical use of the Lokomat training parameters, i.e. guidance, speed and bodyweight support, requires a good understanding of how these parameters affect the neuromuscular control of post-stroke hemiparetic gait. Materials and methods: Ten stroke patients (unilateral paresis, 7 females, 64.5 ± 6.4 years, >3months post-stroke, FAC scores 2–4)) walked in the Lokomat under varying parameter settings: 50% or 100% guidance, 0.28 or 0.56m/s, 0% or 50% bodyweight support. Electromyography was recorded bilaterally from Gluteus Medius, Biceps Femoris, Vastus Lateralis, Medial Gastrocnemius, and Tibialis Anterior. Pressure sensors placed under the feet were used to determine the level of temporal gait symmetry. Results: Varying guidance and bodyweight support had little effect on muscle activity, but increasing treadmill speed led to increased activity in both the affected (Biceps Femoris, Medial Gastrocnemius, Tibialis Anterior) and unaffected leg (all muscles). The level of temporal symmetry was unaffected by the parameter settings. Conclusions: The Lokomat training parameters are generally ineffective in shaping short term muscle activity and step symmetry patients with hemiparetic stroke, as speed is the only parameter that significantly affects muscular amplitude. Trial Registration: d.n.a.IMPLICATIONS FOR REHABILITATIONThe Lokomat is a commercially available gait trainer that can be used for gait rehabilitation in post-stroke hemiparetic patients.This study shows that muscle amplitude is generally low during Lokomat guided walking, and that treadmill Speed is the main training parameter to influence muscular output in stroke patients during Lokomat walking.Varying Guidance and Bodyweight Support within a clinical relevant range barely affected muscle activity, and temporal step symmetry was unaffected by variation in any of the training parameters.Based on the findings it is advised to increase speed as early as possible during Lokomat therapy, or use other means (e.g. feedback or instructions) to stimulate active involvement of patients during training. The Lokomat is a commercially available gait trainer that can be used for gait rehabilitation in post-stroke hemiparetic patients. This study shows that muscle amplitude is generally low during Lokomat guided walking, and that treadmill Speed is the main training parameter to influence muscular output in stroke patients during Lokomat walking. Varying Guidance and Bodyweight Support within a clinical relevant range barely affected muscle activity, and temporal step symmetry was unaffected by variation in any of the training parameters. Based on the findings it is advised to increase speed as early as possible during Lokomat therapy, or use other means (e.g. feedback or instructions) to stimulate active involvement of patients during training.</p

Справа Івана Дзюби

У статті автор, використовуючи документи Галузевого державного архіву СБ України, досліджує постать видатного літературознавця, громадського діяча Івана Дзюби у контексті боротьби співробітників органів держбезпеки УРСР з «українським буржуазним націоналізмом».В статье автор, используя документы Отраслевого государственного архива СБ Украины, исследует личность выдающегося литературоведа, общественного деятеля Ивана Дзюбы в контексте борьбы сотрудников органов госбезопасности УССР с «украинским буржуазным национализмом».Using the documents of State branch archive of State Security of Ukraine, the author investigates the personality of Ivan Dzyuba during the struggle of KGB of the UkSSR against the «Ukrainian bourgeois nationalism»

Regulated Membrane Localization of Tiam1, Mediated by the NH2-terminal Pleckstrin Homology Domain, Is Required for Rac-dependent Membrane Ruffling and C-Jun NH2-terminal Kinase Activation

Rho-like GTPases, including Cdc42, Rac, and Rho, regulate signaling pathways that control actin cytoskeletal structures and transcriptional activation. The Tiam1 gene encodes an activator of Rac1, and similarly to constitutively activated (V12)Rac1, overexpression of Tiam1 in fibroblasts induces the formation of membrane ruffles. Tiam1 contains a Dbl homology (DH) domain and adjacent pleckstrin homology (PH) domain, hallmarks for activators of Rho-like GTPases. Unique for Tiam1 are an additional PH domain and a Discs-large homology region in the NH2-terminal part of the protein. Here we show that both in fibroblasts and COS cells, membrane localization of Tiam1 is required for the induction of membrane ruffling. A detailed mutational analysis, in combination with confocal laser scanning microscopy and immunoelectron microscopy, demonstrates that the NH2-terminal PH domain of Tiam1, but not the DH-adjacent PH domain, is essential for membrane association. This NH2-terminal PH domain of Tiam1 can be functionally replaced by the myristoylated membrane localization domain of c-Src, indicating that the primary function of this PH domain is to localize the protein at the membrane. After serum starvation, both membrane association of Tiam1 and ruffling can be induced by serum, suggesting that receptor stimulation induces membrane translocation of Tiam1. Similar to V12Rac1, Tiam1 stimulates the activity of the c-Jun NH2-terminal kinase (JNK). This Rac-dependent stimulation of JNK also requires membrane association of Tiam1. We conclude that the regulated membrane localization of Tiam1 through its NH2-terminal PH domain determines the activation of distinct Rac-mediated signaling pathways

Differences in muscle activity and temporal step parameters between Lokomat guided walking and treadmill walking in post-stroke hemiparetic patients and healthy walkers

Background: The Lokomat is a robotic exoskeleton that can be used to train gait function in hemiparetic stroke. To purposefully employ the Lokomat for training, it is important to understand (1) how Lokomat guided walking affects muscle activity following stroke and how these effects differ between patients and healthy walkers, (2) how abnormalities in the muscle activity of patients are modulated through Lokomat guided gait, and (3) how temporal step characteristics of patients were modulated during Lokomat guided walking. Methods: Ten hemiparetic stroke patients (> 3 months post-stroke) and ten healthy age-matched controls walked on the treadmill and in the Lokomat (guidance force 50%, no bodyweight support) at matched speeds (0.56 m/s). Electromyography was used to record the activity of Gluteus Medius, Biceps Femoris, Vastus Lateralis, Medial Gastrocnemius and Tibialis Anterior, bilaterally in patients and of the dominant leg in healthy walkers. Pressure sensors placed in the footwear were used to determine relative durations of the first double support and the single support phases. Results: Overall, Lokomat guided walking was associated with a general lowering of muscle activity compared to treadmill walking, in patients as well as healthy walkers. The nature of these effects differed between groups for specific muscles, in that reductions in patients were larger if muscles were overly active during treadmill walking (unaffected Biceps Femoris and Gluteus Medius, affected Biceps Femoris and Vastus Lateralis), and smaller if activity was already abnormally low (affected Medial Gastrocnemius). Also, Lokomat guided walking was associated with a decrease in asymmetry in the relative duration of the single support phase. Conclusions: In stroke patients, Lokomat guided walking results in a general reduction of muscle activity, that affects epochs of overactivity and epochs of reduced activity in a similar fashion. These findings should be taken into account when considering the clinical potential of the Lokomat training environment in stroke, and may inform further developments in the design of robotic gait trainers

The Rac activator Tiam1 is required for α3β1-mediated laminin-5 deposition, cell spreading, and cell migration

The Rho-like guanosine triphosphatase Rac1 regulates various signaling pathways, including integrin-mediated adhesion and migration of cells. However, the mechanisms by which integrins signal toward Rac are poorly understood. We show that the Rac-specific guanine nucleotide exchange factor Tiam1 (T-lymphoma invasion and metastasis 1) is required for the integrin-mediated laminin (LN)-5 deposition, spreading, and migration of keratinocytes. In contrast to wild-type keratinocytes, Tiam1-deficient (Tiam1−/−) keratinocytes are unable to adhere to and spread on a glass substrate because they are unable to deposit their own LN5 substrate. Both Tiam1 and V12Rac1 can rescue the defects of Tiam1−/− keratinocytes, indicating that these deficiencies are caused by impaired Tiam1-mediated Rac activation. Tiam1−/− cells are unable to activate Rac upon α3β1-mediated adhesion to an exogenous LN5 substrate. Moreover, Tiam1 deficiency impairs keratinocyte migration in vitro and reepithelialization of excision wounds in mouse skin. Our studies indicate that Tiam1 is a key molecule in α3β1-mediated activation of Rac, which is essential for proper production and secretion of LN5, a requirement for the spreading and migration of keratinocytes