Focal Adhesion Kinases in Adhesion Structures and Disease

Cell adhesion to the extracellular matrix (ECM) is essential for cell migration, proliferation, and embryonic development. Cells can contact the ECM through a wide range of matrix contact structures such as focal adhesions, podosomes, and invadopodia. Although they are different in structural design and basic function, they share common remodeling proteins such as integrins, talin, paxillin, and the tyrosine kinases FAK, Pyk2, and Src. In this paper, we compare and contrast the basic organization and role of focal adhesions, podosomes, and invadopodia in different cells. In addition, we discuss the role of the tyrosine kinases, FAK, Pyk2, and Src, which are critical for the function of the different adhesion structures. Finally, we discuss the essential role of these tyrosine kinases from the perspective of human diseases

Bone Resorption by Osteoclasts: Molecular Mechanism of Pyk2 dephosphorylation by Dynamin

poster abstractOsteoporosis is a bone disease that affects hundreds of millions of people worldwide and is characterized by low bone mass and structural deterioration of bone tissue which increases the risk of bone fracture, frailty, morbidity and mortality. Excessive bone loss is caused by osteoclasts which degrade the organic and inorganic components of bone. The specific aim of this study is to identify and characterize the signaling proteins in osteoclasts that are responsible for the bone resorbing activity of these cells. The non-receptor tyrosine kinase, Pyk2, is highly expressed in osteoclasts. Mice lacking Pyk2 have an increase in bone mass due to impairment in osteoclast function. It has been demonstrated that phosphorylation of Pyk2 at Y402 is very important for osteoclast spreading and bone resorption. Our group also reported that the GTPase dynamin controls osteoclast bone resorption in part by leading to the dephosphorylation of Pyk2, thus decreasing Pyk2’s kinase activity. In the current study we examined the intracellular mechanism by which dynamin regulates Pyk2 dephosphorylation. Our findings demonstrated that Pyk2 dephosphorylation is predominately due to GTPase activity of dynamin since expression of dynamin mutants that have reduced affinity for GTP or exhibit defective GTPase activity resulted in an increase in Pyk2 Y402 phosphorylation. We also found that that Pyk2 phosphorylation was rescued in the presence of phenyl arsine oxide (PAO), a chemical inhibitor of tyrosine phosphatases and our preliminary results indicate that the tyrosine phosphatase PTP-PEST is involved in the dynamin-mediated dephosphorylation of Pyk2. Understanding the intracellular mechanism that regulates osteoclast function may lead to the identification of novel proteins that can be targeted by anti-resorptive therapies to treat bone related diseases. Over the past few decades, bisphosphonates have played a significant role in the treatment of osteoporosis. Unfortunately, osteonecrosis of the jaw has been recently described as a harmful side effect of bisphosphonate therapy, emphasizing the need to develop alternative approaches to treat osteoporosis. Novel therapeutic approaches may one day involve inhibitors to tyrosine kinases such as Pyk2 or involve combination therapies where inhibitors are paired with bisphosphonates as a way to boost the efficacy of anti-resorptive therapies with fewer side-effects

An aging evaluation of the bearing performances of glass fiber composite laminate in salt spray fog environment

The aim of the present paper is to assess the bearing performance evolution of pinned, glass-composite laminates due to environmental aging in salt-spray fog tests. Glass fibers/epoxy pinned laminates were exposed for up to 60 days in salt-spraying, foggy environmental conditions (according to ASTM B117 standard). In order to evaluate the relationship between mechanical failure mode and joint stability over increasing aging time, different single lap joints, measured by the changing hole diameter (D), laminate width (W) and hole free edge distance (E), were characterized at varying aging steps. Based on this approach, the property-structure relationship of glass-fibers/epoxy laminates was assessed under these critical environmental conditions. Furthermore, an experimental 2D failure map, clustering main failure modes in the plane E/D versus W/D ratios, was generated, and its cluster variation was analyzed at each degree of aging

Dynamin and PTP-PEST cooperatively regulate Pyk2 dephosphorylation in osteoclasts

Bone loss is caused by the dysregulated activity of osteoclasts which degrade the extracellular bone matrix. The tyrosine kinase Pyk2 is highly expressed in osteoclasts, and mice lacking Pyk2 exhibit an increase in bone mass, in part due to impairment of osteoclast function. Pyk2 is activated by phosphorylation at Y402 following integrin activation, but the mechanisms leading to Pyk2 dephosphorylation are poorly understood. In the current study, we examined the mechanism of action of the dynamin GTPase on Pyk2 dephosphorylation. Our studies reveal a novel mechanism for the interaction of Pyk2 with dynamin, which involves the binding of Pyk2's FERM domain with dynamin's plextrin homology domain. In addition, we demonstrate that the dephosphorylation of Pyk2 requires dynamin's GTPase activity and is mediated by the tyrosine phosphatase PTP-PEST. The dephosphorylation of Pyk2 by dynamin and PTP-PEST may be critical for terminating outside-in integrin signaling, and for stabilizing cytoskeletal reorganization during osteoclast bone resorption

Tailored Torsion and Bending-Resistant Avian-Inspired Structures

The escalating demand for torsion- and bending-resistant structures paired with the need for more efficient use of materials and geometries, have led to novel bio-inspired ingenious solutions. However, lessons from Nature could be as inspiring as they are puzzling: plants and animals offer an enormous range of promising but hierarchically complex configurations. Avian bones are prominent candidates for addressing the torsional and bending issue. They present a unique intertwining of simple components: helicoidal ridges and crisscrossing struts, able to bear flexural and twisting actions of winds. Here, it is set how to harmonically move from the natural to the engineering level to formalize and analyze the biological phenomena under controlled design conditions. The effect of ridges and struts is isolated and combined toward tailored torsion and bending-resistant arrangements. Then the biological level is revisited to extrapolate the avian allometric design approach and is translated into multiscale lightweight structures at the engineering level. This study exploits the complexity of Nature and the scalability that characterizes the evolutionary design of bird bones through the design and fabrication versatility allowed by additive manufacturing technologies. This paves the way for exploring the transferability of the proposed solution at multiple engineering scales

Osteoblast differentiation and migration are regulated by Dynamin GTPase activity

Bone formation is controlled by osteoblasts but the signaling proteins that control osteoblast differentiation and function are still unclear. We examined if the dynamin GTPase, which is associated with actin remodeling and migration in other cells, plays a role in osteoblast differentiation and migration. Dynamin mRNA was expressed in primary osteoblasts throughout differentiation (0–21 days). However, alkaline phosphatase (ALP) activity, a marker of osteoblast differentiation, was decreased in osteoblasts over-expressing dynamin. Conversely, ALP activity was increased following shRNA-mediated knockdown of dynamin and in osteoblasts treated with the dynamin inhibitor, dynasore. Dynasore also reduced c-fos and osterix expression, markers of early osteoblasts, suggesting a role for dynamin in pre-osteoblast to osteoblast differentiation. Since dynamin GTPase activity is regulated by tyrosine phosphorylation, we examined the mechanism of dynamin dephosphorylation in osteoblasts. Dynamin formed a protein complex with the tyrosine phosphatase PTP-PEST and inhibition of phosphatase activity increased the level of phosphorylated dynamin. Further, PTP-PEST blocked the Src-mediated increase in the phosphorylation and GTPase activity of wild-type dynamin but not the phosphorylation mutant dynY231F/Y597F. Although ALP activity was increased in osteoblasts expressing GTPase-defective dynK44A, and to a lesser extent dynY231F/Y597F, osteoblast migration was significantly inhibited by dynK44A and dynY231F/Y597F. These studies demonstrate a novel role for dynamin GTPase activity and phosphorylation in osteoblast differentiation and migration, which may be important for bone formation

First-in-man craniectomy and asportation of solitary cerebellar metastasis in COVID-19 patient: A case report

Introduction: Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) outbreak has an impact on the delivery of neurosurgical care, and it is changing the perioperative practice worldwide. We present the first case in the literature of craniectomy procedure and asportation of a solitary cerebellar metastasis of the oesophagus squamous carcinoma in a 77 years old woman COVID-19 positive. In these particular circumstances, we show that adequate healthcare resources and risk assessments are essential in the management of COVID-19 patients referred to emergency surgery. Presentation of case: The case here presented was treated in 2019 for squamous carcinoma of the oesophagus. In April 2020, she presented a deterioration of her clinical picture consisting of dysphagia, abdominal pain, hyposthenia and ataxia. A Head CT scan was performed, which showed the presence of a solitary cerebellar metastasis. Her associated SARS-CoV-2 positivity status represented the principal clinical concern throughout her hospitalisation. Discussion: The patient underwent a suboccipital craniectomy procedure with metastasis asportation. She tested positive for SARS-CoV-2 in the pre- and post-operative phases, but she was not admitted to the intensive care unit because she did not present any respiratory complications. Her vital parameters and inflammation indexes fell within the reference ranges, and she was kept in isolation for 16 days in our neurosurgical unit following strict COVID-19 measures. She was asymptomatic and not treated for any of the specific and non-specific symptoms of COVID-19. Conclusion: This is the first case reported of solitary cerebellar metastasis of oesophagus carcinoma operated on a COVID-19 positive patient. It shows that asymptomatic COVID-19 positive patients can undergo major emergency surgeries without the risk of infecting the operating team if adequate Personal Protection Equipment (PPE) is used. The patient remained asymptomatic and did not develop the disease's active phase despite undergoing a stressful event such as a major emergency neurosurgical procedure. In the current crisis, a prophylactic COVID-19 screening test can identify asymptomatic patients undergoing major emergency surgery and adequate resource planning and Personal Protective Equipment (PPE) for healthcare workers can minimise the effect of the COVID-19 pandemic

Adsorption performance and thermodynamic analysis of SAPO-34 silicone composite foams for adsorption heat pump applications

Abstract In the present work, adsorption performances of an innovative composite adsorber, based on SAPO-34-silicone composite macro-cellular foams, are reported. The choice of a foamed structure was assessed to improve the water vapor access towards the embedded zeolite keeping good adsorption heat pump dynamic performance. Depending on zeolite amount used as filler, zeolite/silicone foams evidenced a soft and open cell configuration (low zeolite content) or rigid and closed one (high zeolite content). Morphological analysis evidenced that the cellular structure of the foam is homogeneous and well distributed along the foam cross section. Adsorption tests showed that the adsorbent foamed samples have very effective adsorption capabilities indicating that the porous structure of the filled pure zeolite was not obstructed. SAPO-34 filler contributed actively, with an efficiency above 90%, to the adsorption performances of the composite foam. Starting from experimental equilibrium data, a simple thermodynamic analysis based on energy balances was carried out for air conditioning application. Results of the analysis demonstrated that foam technology can guarantee cooling COP up to 7% higher than that estimated for the typical adsorber solution based on loose adsorbent grains inside an aluminum finned-flat tube heat exchanger, which is very promising for practical application in adsorption heat pumps

The Rho-GEF Kalirin regulates bone mass and the function of osteoblasts and osteoclasts

Bone homeostasis is maintained by the balance between bone resorption by osteoclasts and bone formation by osteoblasts. Dysregulation in the activity of the bone cells can lead to osteoporosis, a disease characterized by low bone mass and an increase in bone fragility and risk of fracture. Kalirin is a novel GTP-exchange factor protein that has been shown to play a role in cytoskeletal remodeling and dendritic spine formation in neurons. We examined Kalirin expression in skeletal tissue and found that it was expressed in osteoclasts and osteoblasts. Furthermore, micro-CT analyses of the distal femur of global Kalirin knockout (Kal-KO) mice revealed significantly reduced trabecular and cortical bone parameters in Kal-KO mice, compared to WT mice, with significantly reduced bone mass in 8, 14 and 36 week-old female Kal-KO mice. Male mice also exhibited a decrease in bone parameters but not to the level seen in female mice. Histomorphometric analyses also revealed decreased bone formation rate in 14 week-old female Kal-KO mice, as well as decreased osteoblast number/bone surface and increased osteoclast surface/bone surface. Consistent with our in vivo findings, the bone resorbing activity and differentiation of Kal-KO osteoclasts was increased in vitro. Although alkaline phosphatase activity by Kal-KO osteoblasts was increased in vitro, Kal-KO osteoblasts showed decreased mineralizing activity, as well as decreased secretion of OPG, which was inversely correlated with ERK activity. Taken together, our findings suggest that deletion of Kalirin directly affects osteoclast and osteoblast activity, leading to decreased OPG secretion by osteoblasts which is likely to alter the RANKL/OPG ratio and promote osteoclastogenesis. Therefore, Kalirin may play a role in paracrine and/or endocrine signaling events that control skeletal bone remodeling and the maintenance of bone mass

Megakaryocytes Regulate Expression of Pyk2 Isoforms and Caspase-mediated Cleavage of Actin in Osteoblasts

The proliferation and differentiation of osteoblast (OB) precursors are essential for elaborating the bone-forming activity of mature OBs. However, the mechanisms regulating OB proliferation and function are largely unknown. We reported that OB proliferation is enhanced by megakaryocytes (MKs) via a process that is regulated in part by integrin signaling. The tyrosine kinase Pyk2 has been shown to regulate cell proliferation and survival in a variety of cells. Pyk2 is also activated by integrin signaling and regulates actin remodeling in bone-resorbing osteoclasts. In this study, we examined the role of Pyk2 and actin in the MK-mediated increase in OB proliferation. Calvarial OBs were cultured in the presence of MKs for various times, and Pyk2 signaling cascades in OBs were examined by Western blotting, subcellular fractionation, and microscopy. We found that MKs regulate the temporal expression of Pyk2 and its subcellular localization. We also found that MKs regulate the expression of two alternatively spliced isoforms of Pyk2 in OBs, which may regulate OB differentiation and proliferation. MKs also induced cytoskeletal reorganization in OBs, which was associated with the caspase-mediated cleavage of actin, an increase in focal adhesions, and the formation of apical membrane ruffles. Moreover, BrdU incorporation in MK-stimulated OBs was blocked by the actin-polymerizing agent, jasplakinolide. Collectively, our studies reveal that Pyk2 and actin play an important role in MK-regulated signaling cascades that control OB proliferation and may be important for therapeutic interventions aimed at increasing bone formation in metabolic diseases of the skeleton