Predominance of Th2 response in human abdominal aortic aneurysm: mistaken identity for IL-4-producing NK and NKT cells?

Abdominal aortic aneurysm (AAA) is a complex remodeling process that involves both synthesis and degradation of extracellular matrix proteins in the aortic wall, leading to decreased tensile strength, progressive dilation and eventual rupture. Chronic inflammation, increased local production of elastin-degrading proteases by inflammatory cells and destruction of medial elastic lamellae play important roles in aneurysm progression. Neovascularization in all layers of the arterial wall is prominent and angiogenesis can facilitate chronic inflammation. It is still unclear what initiates aneurysmal dilation and what determines its progression. The complex nature of the process has defied elucidation. Apart from macrophages, the predominant immune cell infiltrates reported so far are CD3(+)T cells that express CD4 and CD8. Infiltrates of type 2 Th cells and their production of IL-4 and IL-5 have been implicated in AAA development. However, NKT and NK cells have a Th0 cytokine profile and can also produce type 2 as well as type 1 (IL-2 and IFNgamma) cytokines. We have demonstrated the presence of NK and NKT cells in AAA tissue. With their growing importance in autoimmunity and transplantation, they may play a role in AAA development. Therefore, there is a need to use a combination of T and NK markers to fully characterize both innate and adaptive lymphoid cell subsets in local inflammatory infiltrates in order to elucidate their roles in AAA progression.     

Integration of molecular dynamics simulation and hotspot residues grafting for de novo scFv design against Salmonella Typhi TolC protein

With the development of de novo binders for protein targets from non‐related scaffolds, many possibilities for therapeutics and diagnostics have been created. In this study, we described the use of de novo design approach to create single‐chain fragment variable (scFv) for Salmonella enterica subspecies enterica serovar Typhi TolC protein. Typhoid fever is a global health concern in developing and underdeveloped countries. Rapid typhoid diagnostics will improve disease management and therapy. In this work, molecular dynamics simulation was first performed on a homology model of TolC protein in POPE membrane bilayer to obtain the central structure that was subsequently used as the target for scFv design. Potential hotspot residues capable of anchoring the binders to the target were identified by docking “disembodied” amino acid residues against TolC surface. Next, scFv scaffolds were selected from Protein Data Bank to harbor the computed hotspot residues. The hotspot residues were then incorporated into the scFv scaffold complementarity determining regions. The designs recapitulated binding energy, shape complementarity, and interface surface area of natural protein‐antibody interfaces. This approach has yielded 5 designs with high binding affinity against TolC that may be beneficial for the future development of antigen‐based detection agents for typhoid diagnostics.     

Aged‐senescent cells contribute to impaired heart regeneration

Aging leads to increased cellular senescence and is associated with decreased potency of tissue‐specific stem/progenitor cells. Here, we have done an extensive analysis of cardiac progenitor cells (CPCs) isolated from human subjects with cardiovascular disease, aged 32–86 years. In aged subjects (>70 years old), over half of CPCs are senescent (p16^INK4A, SA‐β‐gal, DNA damage γH2AX, telomere length, senescence‐associated secretory phenotype [SASP]), unable to replicate, differentiate, regenerate or restore cardiac function following transplantation into the infarcted heart. SASP factors secreted by senescent CPCs renders otherwise healthy CPCs to senescence. Elimination of senescent CPCs using senolytics abrogates the SASP and its debilitative effect in vitro. Global elimination of senescent cells in aged mice (INK‐ATTAC or wild‐type mice treated with D + Q senolytics) in vivo activates resident CPCs and increased the number of small Ki67‐, EdU‐positive cardiomyocytes. Therapeutic approaches that eliminate senescent cells may alleviate cardiac deterioration with aging and restore the regenerative capacity of the heart.     

Establishing Restoration Strategy of Eastern Oyster via a Coupled Biophysical Transport Model

For marine fish and invertebrates, larval dispersal plays a critical role in determining connections among source and sink habitats, and the lack of a predictive understanding of larval dispersal is a fundamental obstacle to the development of spatially explicit restoration plans for marine populations. We investigated larval dispersal patterns of eastern oyster in an estuary along the Northern Gulf of Mexico under different simulation scenarios of tidal amplitude and phase, river discharge, wind direction, and larval vertical migration, using a coupled biophysical transport model. We focused on the dispersal of larvae released from the commercially exploited (Cedar Point, CP) and non‐exploited (Bon Secour Bay, BSB) oyster populations. We found that high flushing rates through the dominant inlet prevented larval exchange between the commercially exploited and non‐exploited populations, resulting in negligible connectivity between them. Variations in tidal amplitude, river discharge and wind direction played a more important role in the amount of larvae retained in Mobile Bay when they are released from CP than from BSB. Under most of the scenarios, larvae from BSB were retained around the spawning area, while larvae from CP showed a predominant westward flow. Net sinking behavior of late‐stage larvae increased larval retention in the bay, but physical transport showed a higher impact in the amount of larvae retained. These findings have enhanced our understanding of larval dispersal of eastern oyster in a wide, shallow estuarine system, and been used to establish spatially explicit strategies for oyster restoration in the Mobile Bay system, Alabama.     

Cadmium affects focal adhesion kinase (FAK) in mesangial cells: Involvement of CaMK‐II and the actin cytoskeleton

The toxic metal ion cadmium (Cd²⁺) induces pleiotropic effects on cell death and survival, in part through effects on cell signaling mechanisms and cytoskeletal dynamics. Linking these phenomena appears to be calmodulin‐dependent activation of the Ca²⁺/calmodulin‐dependent protein kinase II (CaMK‐II). Here we show that interference with the dynamics of the filamentous actin cytoskeleton, either by stabilization or destabilization, results in disruption of focal adhesions at the ends of organized actin structures, and in particular the loss of vinculin and focal adhesion kinase (FAK) from the contacts is a result. Low‐level exposure of renal mesangial cells to CdCl₂ disrupts the actin cytoskeleton and recapitulates the effects of manipulation of cytoskeletal dynamics with biological agents. Specifically, Cd²⁺ treatment causes loss of vinculin and FAK from focal contacts, concomitant with cytoskeletal disruption, and preservation of cytoskeletal integrity with either a calmodulin antagonist or a CaMK‐II inhibitor abrogates these effects of Cd²⁺. Notably, inhibition of CaMK‐II decreases the migration of FAK‐phosphoTyr925 to a membrane‐associated compartment where it is otherwise sequestered from focal adhesions in a Cd²⁺‐dependent manner. These results add further insight into the mechanism of the CaMK‐II‐dependent effects of Cd²⁺ on cellular function. J. Cell. Biochem. 114: 1832–1842, 2013. © 2013 Wiley Periodicals, Inc.     

Gliosis in the Mice Hippocampus Without Neuronal Death After Systemic Administration of High Dosage of Tetanus Toxin

Tetanus toxin (TeT), an exotoxin, has been studied to cause tetanus in mammalian brains, and it can block the release of some neurotransmitters and affect seizure propagation. In the present study, we investigated neuronal damage/death and glial changes in the mouse hippocampus after systemic administration (intraperitoneal injection) of TeT 10 and 100 ng/kg. In both the 10 and 100 ng/kg TeT-treated groups, no neuronal death occurred in any subregions of the mouse hippocampus until 24 h post-treatment; however, there were changes in glia in the hippocampus depending on time course and dosage. The morphology of GFAP-immunoreactive astrocytes and Iba-1-immunoreactive microglia was apparently changed in the 100 ng/kg TeT treated-group compared to the 10 ng/kg TeT treated-group. In the 100 ng/kg TeT treated-group, they were increased in size and their immunoreactivity was distinctively increased from 12 h post-treatment. We also found that their protein levels were increased in the hippocampus at 12 h post-treatment of 100 ng/kg TeT. In conclusion, these results indicate that the systemic administration of 100 ng/kg TeT induced a distinctive microglia changes in the mouse hippocampus without any neuronal death/damage.     

Ginkgolide B production in cultured cells derived from Ginkgo biloba L. leaves

Callus cultures and cell suspension cultures derived from Ginkgo biloba L. leaves produced ginkgolidc B. In cell suspension cultures, the production reached a maximum by the 13th day of subculture and followed by a sharp decrease. The medium of Murashige and Skoog induced the highest ginkgolide B content in cultures while the medium of Schenk and Hildebrandt promoted cell growth. For the maximal production of ginkgolide B, cells were cultured in Murashige and Skoog medium modified to contain 1.0 mg/l of -naphthaleneacetic acid, 0.1 mg/1 of kinetin, 30 g/1 sucrose and 1.25 mM potassium phosphate with a molar ratio of ammonium to nitrate ions of 1 3.Abbreviations B5 Gamborg et al (1968) medium - GKB Ginkgolide B - MS Murashige and Skoog (1962) medium - NAA -naphthaleneacetic aicd - SH Schenk and Hildebrandt (1972) medium     

EphA2 Mutation in Lung Squamous Cell Carcinoma Promotes Increased Cell Survival,Cell Invasion,Focal Adhesions,and Mammalian Target of Rapamycin Activation

Non-small cell lung cancer (NSCLC) has a poor prognosis and improved therapies are needed. Expression of EphA2 is increased in NSCLC metastases. In this study, we investigated EphA2 mutations in NSCLC and examined molecular pathways involved in NSCLC. Tumor and cell line DNA was sequenced. One EphA2 mutation was modeled by expression in BEAS2B cells, and functional and biochemical studies were conducted. A G391R mutation was detected in H2170 and 2/28 squamous cell carcinoma patient samples. EphA2 G391R caused constitutive activation of EphA2 with increased phosphorylation of Src, cortactin, and p130^Cas. Wild-type (WT) and G391R cells had 20 and 40% increased invasiveness; this was attenuated with knockdown of Src, cortactin, or p130^Cas. WT and G391R cells demonstrated a 70% increase in focal adhesion area. Mammalian target of rapamycin (mTOR) phosphorylation was increased in G391R cells with increased survival (55%) compared with WT (30%) and had increased sensitivity to rapamycin. A recurrent EphA2 mutation is present in lung squamous cell carcinoma and increases tumor invasion and survival through activation of focal adhesions and actin cytoskeletal regulatory proteins as well as mTOR. Further study of EphA2 as a therapeutic target is warranted.     

The diabetic rat kidney mediates inosituria and selective urinary partitioning of D-chiro-inositol

Diabetic nephropathy is a serious complication of diabetes mellitus with a pressing need for effective metabolic markers to detect renal impairment. Of potential significance are the inositol compounds, myo-inositol (MI), and the less abundant stereoisomer, D-chiro-inositol (DCI), which are excreted at increased levels in the urine in diabetes mellitus, a phenomenon known as inosituria. There is also a selective urinary excretion of DCI compared to MI. As the biological origins of altered inositol metabolism in diabetes mellitus are unknown, the aim of this study was to determine whether the diabetic kidney was directly responsible. Kidneys isolated from four-week streptozotocin-induced diabetic rats were characterized by a 3-fold reduction in glomerular filtration rate (GFR) compared to matched non-diabetic kidneys. When perfused with fixed quantities of MI (50 µM) and DCI (5 µM) under normoglycemic conditions (5 mM glucose), GFR-normalized urinary excretion of MI was increased by 1.7-fold in diabetic vs. non-diabetic kidneys. By comparison, GFR-normalized urinary excretion of DCI was increased by 4-fold. Perfusion conditions replicating hyperglycemia (20 mM glucose) potentiated DCI but not MI urinary excretion in both non-diabetic and diabetic kidneys. Overall, there was a 2.4-fold increase in DCI urinary excretion compared to MI in diabetic kidneys that was independent of glucose ambience. This increased urinary excretion of DCI and MI in diabetic kidneys occurred despite increased renal expression of the inositol transporters, sodium myo-inositol transporter subtype 1 and 2 (SMIT1 and SMIT2). These findings show that the diabetic kidney primarily mediates inosituria and altered urinary partitioning of MI and DCI. Urinary inositol levels might therefore serve as an indicator of impaired renal function in diabetes mellitus with wider implications for monitoring chronic kidney disease.     

Photoluminescence imaging of europium (III)‐doped γ‐Al2O3 nanofiber structures

Aluminium oxide (Al₂O₃) has widely been used for catalysts, insulators, and composite materials for diverse applications. Herein, we demonstrated if γ‐Al₂O₃ was useful as a luminescence support material for europium (Eu) (III) activator ion. The hydrothermal method and post‐thermal treatment at 800°C were employed to synthesize Eu(III)‐doped γ‐Al₂O₃ nanofibre structures. Luminescence characteristics of Eu(III) ions in Al₂O₃ matrix were fully understood by taking 2D and 3D‐photoluminescence imaging profiles. Various sharp emissions between 580 to 720 nm were assigned to the ⁵D₀→⁷F_J (J = 0, 1, 2, 3, 4) transitions of Eu(III) activators. On the basis of X‐ray diffraction crystallography, Auger elemental mapping and the asymmetry ratio, Eu(III) ions were found to be well doped into the γ‐Al₂O₃ matrix at a low (1 mol%) doping level. A broad emission at 460 nm was substantially increased upon higher (2 mol%) Eu(III) doping due to defect creation. The first 3D photoluminescence imaging profiles highlight detailed understanding of emission characteristics of Eu(III) ions in Al oxide‐based phosphor materials and their potential applications.