Lithium Treatment of APPSwDI/NOS2−/− Mice Leads to Reduced Hyperphosphorylated Tau, Increased Amyloid Deposition and Altered Inflammatory Phenotype

Lithium is an anti-psychotic that has been shown to prevent the hyperphosphorylation of tau protein through the inhibition of glycogen-synthase kinase 3-beta (GSK3β). We recently developed a mouse model that progresses from amyloid pathology to tau pathology and neurodegeneration due to the genetic deletion of NOS2 in an APP transgenic mouse; the APPSwDI/NOS2−/− mouse. Because this mouse develops tau pathology, amyloid pathology and neuronal loss we were interested in the effect anti-tau therapy would have on amyloid pathology, learning and memory. We administered lithium in the diets of APPSwDI/NOS2−/− mice for a period of eight months, followed by water maze testing at 12 months of age, immediately prior to sacrifice. We found that lithium significantly lowered hyperphosphorylated tau levels as measured by Western blot and immunocytochemistry. However, we found no apparent neuroprotection, no effect on spatial memory deficits and an increase in histological amyloid deposition. Aβ levels measured biochemically were unaltered. We also found that lithium significantly altered the neuroinflammatory phenotype of the brain, resulting in enhanced alternative inflammatory response while concurrently lowering the classical inflammatory response. Our data suggest that lithium may be beneficial for the treatment of tauopathies but may not be beneficial for the treatment of Alzheimer's disease

Effect of Lactobacillus acidophilus supernatants on body weight and leptin expression in rats

<p>Abstract</p> <p>Background</p> <p><it>Lactobacillus </it>extracts and supernatants have been used as probiotics in human and veterinary medicine for their ability to enhance wound healing and immunity. Previous data from our laboratory demonstrated that <it>Lactobacillus </it>supernatant (LS) stimulated wound healing, angiogenesis and proliferation of embryonic cells after topical application. This current study shows that LS after its administration into the cerebral ventricles of male rats exerts systemic effects.</p> <p>Methods</p> <p>The right lateral cerebral ventricle of young male rats was accessed through intracerebroventricular cannulation (ICV) under anesthesia and aseptic conditions. One group of control rats received saline solution, a second control group received 0.8 M lactic acid solution (to control for acidity of LS), and a third group received LS. The animals were sacrificed 12, 24, 48, 96 and 120 hours after the injection. Selected tissues were collected, fixed in 10% buffered formalin and used for immunohistochemistry and <it>in situ </it>hybridization. Other tissues were frozen and extracted for immunoblotting</p> <p>Results</p> <p>LS-injected animals had a slight decrease in body weight when compared to their initial weight and to both control groups. Using immunohistochemistry and <it>in situ </it>hybridization leptin expression was studied in multiple brain sections and peripheral adipose tissue of control and LS-injected rats. Strong cytoplasmic stain was observed by both techniques in neurons of the cerebral cortex, thalamus, hypothalamus, hippocampus and, to lesser degree, in the cells of the choroid plexus in the LS-injected rats. Control animals demonstrated much less intense staining in neurons located in the same regions using immunohistochemistry and almost no staining with <it>in situ </it>hybridization technique. Adipose tissue exhibited slight presence of leptin in LS-treated animals. In contrast no immunohistochemical staining for GM-CSF and TNFα was observed in brains from control and treated rats. Western blotting showed mild increase in leptin and leptin receptors in intestines and retroperitoneal adipose tissues of LS-injected rats.</p> <p>Conclusion</p> <p>This study demonstrates that direct administration of LS into rat CNS leads to a decrease in body weight of rats and an increase in the expression of leptin in specific areas of the brain and retroperitoneal adipose tissue.</p

Molecular imaging of glycan chains couples cell-wall polysaccharide architecture to bacterial cell

Biopolymer composite cell walls maintain cell shape and resist forces in plants, fungi and bacteria. Peptidoglycan, a crucial antibiotic target and immunomodulator, performs this role in bacteria. The textbook structural model of peptidoglycan is a highly ordered, crystalline material. Here we use atomic force microscopy (AFM) to image individual glycan chains in peptidoglycan from Escherichia coli in unprecedented detail. We quantify and map the extent to which chains are oriented in a similar direction (orientational order), showing it is much less ordered than previously depicted. Combining AFM with size exclusion chromatography, we reveal glycan chains up to 200 nm long. We show that altered cell shape is associated with substantial changes in peptidoglycan biophysical properties. Glycans from E. coli in its normal rod shape are long and circumferentially oriented, but when a spheroid shape is induced (chemically or genetically) glycans become short and disordered

Expression and prognostic significance of THBS1, Cyr61 and CTGF in esophageal squamous cell carcinoma

<p>Abstract</p> <p>Background</p> <p>Thrombospondin1 (THBS1), cystene-rich protein 61 (Cyr61) and connective tissue growth factor (CTGF) are all involved in the transforming growth factor-beta (TGF-β) signal pathway, which plays an important role in the tumorigenesis. The purpose of this study is to explore the expression and prognostic significance of these proteins in esophageal squamous cell carcinoma (ESCC).</p> <p>Methods</p> <p>We used immunohistochemistry and western blotting to examine the expression status of THBS1, Cyr61 and CTGF in ESCC. Correlations of THBS1, Cyr61 and CTGF over-expressions with various clinicopathologic factors were also determined by using the Chi-square test or Fisher's exact probability test. Survival analysis was assessed by the Kaplan-Meier analysis and the log-rank test. Relative risk was evaluated by the multivariate Cox proportional hazards model.</p> <p>Results</p> <p>THBS1, Cyr61 and CTGF were all over-expressed in ESCC. THBS1 over-expression was significantly associated with TNM stage (<it>P </it>= 0.029) and regional lymph node involvement (<it>P </it>= 0.026). Kaplan-Meier survival analysis showed that over-expression of THBS1, Cyr61 or CTGF was related to poor survival of ESCC patients (<it>P </it>= 0.042, <it>P </it>= 0.020, <it>P </it>= 0.018, respectively). Multivariate Cox analysis demonstrated that Cyr61 and CTGF were independent factors in prognosis of ESCC.</p> <p>Conclusion</p> <p>Cyr61, CTGF and THBS1 were all over-expressed in ESCC and might be new molecular markers to predict the prognosis of ESCC patients.</p

Calculations of binding affinity between C8-substituted GTP analogs and the bacterial cell-division protein FtsZ

The FtsZ protein is a self-polymerizing GTPase that plays a central role in bacterial cell division. Several C8-substituted GTP analogs are known to inhibit the polymerization of FtsZ by competing for the same binding site as its endogenous activating ligand GTP. Free energy calculations of the relative binding affinities to FtsZ for a set of five C8-substituted GTP analogs were performed. The calculated values agree well with the available experimental data, and the main contribution to the free energy differences is determined to be the conformational restriction of the ligands. The dihedral angle distributions around the glycosidic bond of these compounds in water are known to vary considerably depending on the physicochemical properties of the substituent at C8. However, within the FtsZ protein, this substitution has a negligible influence on the dihedral angle distributions, which fall within the narrow range of −140° to −90° for all investigated compounds. The corresponding ensemble average of the coupling constants 3J(C4,H1′) is calculated to be 2.95 ± 0.1 Hz. The contribution of the conformational selection of the GTP analogs upon binding was quantified from the corresponding populations. The obtained restraining free energy values follow the same trend as the relative binding affinities to FtsZ, indicating their dominant contribution

Expression of CCN family of genes in human skin in vivo and alterations by solar-simulated ultraviolet irradiation

The CCN family of proteins is involved in diverse biological functions such as cell growth, adhesion, migration, angiogenesis, and regulation of extracellular matrix. We have investigated expression of CCN family genes and alternations induced by solar-simulated ultraviolet irradiation in human skin in vivo. Transcripts of all six CCN genes were expressed in human skin in vivo. CCN5 was most abundantly expressed followed by CCN2>CCN3>CCN1>CCN4>CCN6. Solar-simulated ultraviolet irradiation increased mRNA expression of CCN1 and CCN2. In contrast, mRNA levels of CCN3, CCN4, CCN5, and CCN6, were reduced. Knowledge gained from this study provides the foundation to explore the functional roles of CCN gene products in cutaneous biology and responses to solar ultraviolet irradiation

Characterization of Modular Bacteriophage Endolysins from Myoviridae Phages OBP, 201ϕ2-1 and PVP-SE1

Peptidoglycan lytic enzymes (endolysins) induce bacterial host cell lysis in the late phase of the lytic bacteriophage replication cycle. Endolysins OBPgp279 (from Pseudomonas fluorescens phage OBP), PVP-SE1gp146 (Salmonella enterica serovar Enteritidis phage PVP-SE1) and 201ϕ2-1gp229 (Pseudomonas chlororaphis phage 201ϕ2-1) all possess a modular structure with an N-terminal cell wall binding domain and a C-terminal catalytic domain, a unique property for endolysins with a Gram-negative background. All three modular endolysins showed strong muralytic activity on the peptidoglycan of a broad range of Gram-negative bacteria, partly due to the presence of the cell wall binding domain. In the case of PVP-SE1gp146, this domain shows a binding affinity for Salmonella peptidoglycan that falls within the range of typical cell adhesion molecules (Kaff = 1.26×106 M−1). Remarkably, PVP-SE1gp146 turns out to be thermoresistant up to temperatures of 90°C, making it a potential candidate as antibacterial component in hurdle technology for food preservation. OBPgp279, on the other hand, is suggested to intrinsically destabilize the outer membrane of Pseudomonas species, thereby gaining access to their peptidoglycan and exerts an antibacterial activity of 1 logarithmic unit reduction. Addition of 0.5 mM EDTA significantly increases the antibacterial activity of the three modular endolysins up to 2–3 logarithmic units reduction. This research work offers perspectives towards elucidation of the structural differences explaining the unique biochemical and antibacterial properties of OBPgp279, PVP-SE1gp146 and 201ϕ2-1gp229. Furthermore, these endolysins extensively enlarge the pool of potential antibacterial compounds used against multi-drug resistant Gram-negative bacterial infections

CCN2 Is Required for the TGF-β Induced Activation of Smad1 - Erk1/2 Signaling Network

Connective tissue growth factor (CCN2) is a multifunctional matricellular protein, which is frequently overexpressed during organ fibrosis. CCN2 is a mediator of the pro-fibrotic effects of TGF-β in cultured cells, but the specific function of CCN2 in the fibrotic process has not been elucidated. In this study we characterized the CCN2-dependent signaling pathways that are required for the TGF-β induced fibrogenic response. By depleting endogenous CCN2 we show that CCN2 is indispensable for the TGF-β-induced phosphorylation of Smad1 and Erk1/2, but it is unnecessary for the activation of Smad3. TGF-β stimulation triggered formation of the CCN2/β3 integrin protein complexes and activation of Src signaling. Furthermore, we demonstrated that signaling through the αvβ3 integrin receptor and Src was required for the TGF-β induced Smad1 phosphorylation. Recombinant CCN2 activated Src and Erk1/2 signaling, and induced phosphorylation of Fli1, but was unable to stimulate Smad1 or Smad3 phosphorylation. Additional experiments were performed to investigate the role of CCN2 in collagen production. Consistent with the previous studies, blockade of CCN2 abrogated TGF-β-induced collagen mRNA and protein levels. Recombinant CCN2 potently stimulated collagen mRNA levels and upregulated activity of the COL1A2 promoter, however CCN2 was a weak inducer of collagen protein levels. CCN2 stimulation of collagen was dose-dependent with the lower doses (<50 ng/ml) having a stimulatory effect and higher doses having an inhibitory effect on collagen gene expression. In conclusion, our study defines a novel CCN2/αvβ3 integrin/Src/Smad1 axis that contributes to the pro-fibrotic TGF-β signaling and suggests that blockade of this pathway may be beneficial for the treatment of fibrosis

DW-MRI as a Biomarker to Compare Therapeutic Outcomes in Radiotherapy Regimens Incorporating Temozolomide or Gemcitabine in Glioblastoma

The effectiveness of the radiosensitizer gemcitabine (GEM) was evaluated in a mouse glioma along with the imaging biomarker diffusion-weighted magnetic resonance imaging (DW-MRI) for early detection of treatment effects. A genetically engineered murine GBM model [Ink4a-Arf−/− PtenloxP/loxP/Ntv-a RCAS/PDGF(+)/Cre(+)] was treated with gemcitabine (GEM), temozolomide (TMZ) +/− ionizing radiation (IR). Therapeutic efficacy was quantified by contrast-enhanced MRI and DW-MRI for growth rate and tumor cellularity, respectively. Mice treated with GEM, TMZ and radiation showed a significant reduction in growth rates as early as three days post-treatment initiation. Both combination treatments (GEM/IR and TMZ/IR) resulted in improved survival over single therapies. Tumor diffusion values increased prior to detectable changes in tumor volume growth rates following administration of therapies. Concomitant GEM/IR and TMZ/IR was active and well tolerated in this GBM model and similarly prolonged median survival of tumor bearing mice. DW-MRI provided early changes to radiosensitization treatment warranting evaluation of this imaging biomarker in clinical trials