Stimulation and oxidative catalytic inactivation of thermolysin by copper•Cys-Gly-His-Lys

[Cu²⁺•Cys-Gly-His-Lys] stimulates thermolysin (TLN) activity at low concentration (below 10 μM) and inhibits the enzyme at higher concentration, with binding affinities of 2.0 and 4.9 μM, respectively. The metal-free Cys-Gly-His-Lys peptide also stimulates TLN activity, with an apparent binding affinity of 2.2 μM. Coordination of copper through deprotonated imine nitrogens, the histidyl nitrogen, and the free N-terminal amino group is consistent with the characteristic absorption spectrum of a Cu²⁺–amino-terminal copper and nickel binding motif (λ _max ∼ 525 nm). The lack of thiol coordination is suggested by both the absence of a thiol to Cu²⁺ charge transfer band and electrochemical studies, since the electrode potential (vs. Ag/AgCl) 0.84 V (ΔE = 92 mV) for the Cu^3+/2+ redox couple obtained for [Cu²⁺•Cys-Gly-His-Lys] was found to be in close agreement with that of a related complex [Cu²⁺•Lys-Gly-His-Lys]⁺ (0.84 V, ΔE = 114 mV). The N-terminal cysteine appears to be available as a zinc-anchoring residue and plays a critical functional role since the [Cu²⁺•Lys-Gly-His-Lys]⁺ homologue exhibits neither stimulation nor inhibition of TLN. Under oxidizing conditions (ascorbate/O₂) the catalyst is shown to mediate the complete irreversible inactivation of TLN at concentrations where enzyme activity would otherwise be stimulated. The observed rate constant for inactivation of TLN activity was determined as k _obs = 7.7 × 10⁻² min⁻¹, yielding a second-order rate constant of (7.7 ± 0.9) × 10⁴ M⁻¹ min⁻¹. Copper peptide mediated generation of reactive oxygen species that subsequently modify active-site residues is the most likely pathway for inactivation of TLN rather than cleavage of the peptide backbone.     

YAP1 increases organ size and expands undifferentiated progenitor cells

The mechanisms that regulate mammalian organ size are poorly understood. It is unclear whether the pathways that control organ size also impinge on stem/progenitor cells. A highly expressed gene in stem cells is YAP1, the ortholog of Drosophila Yorkie, a downstream component of the Hippo pathway. Mutations in components of this pathway produce tissue overgrowth phenotypes in the fly whereas mammalian orthologs, like salvador, merlin, LATS, and YAP1, have been implicated in tumorigenesis. We report here that YAP1 increases organ size and causes aberrant tissue expansion in mice. YAP1 activation reversibly increases liver size more than 4-fold. In the intestine, expression of endogenous YAP1 is restricted to the progenitor/stem cell compartment, and activation of YAP1 expands multipotent undifferentiated progenitor cells, which differentiate upon cessation of YAP1 expression. YAP1 stimulates Notch signaling, and administration of gamma-secretase inhibitors suppressed the intestinal dysplasia caused by YAP1. Human colorectal cancers expressing higher levels of YAP1 share molecular aspects with YAP1-induced dysplastic growth in the mouse. Our data show that the Hippo signaling pathway regulates organ size in mammals and can act on stem cell compartments, indicating a potential link between stem/progenitor cells, organ size, and cancer.     

Characterization of human embryonic stem cell lines by the International Stem Cell Initiative

The International Stem Cell Initiative characterized 59 human embryonic stem cell lines from 17 laboratories worldwide. Despite diverse genotypes and different techniques used for derivation and maintenance, all lines exhibited similar expression patterns for several markers of human embryonic stem cells. They expressed the glycolipid antigens SSEA3 and SSEA4, the keratan sulfate antigens TRA-1-60, TRA-1-81, GCTM2 and GCT343, and the protein antigens CD9, Thy1 (also known as CD90), tissue-nonspecific alkaline phosphatase and class 1 HLA, as well as the strongly developmentally regulated genes NANOG, POU5F1 (formerly known as OCT4), TDGF1, DNMT3B, GABRB3 and GDF3. Nevertheless, the lines were not identical: differences in expression of several lineage markers were evident, and several imprinted genes showed generally similar allele-specific expression patterns, but some gene-dependent variation was observed. Also, some female lines expressed readily detectable levels of XIST whereas others did not. No significant contamination of the lines with mycoplasma, bacteria or cytopathic viruses was detected.     

Chronic Inflammation Permanently Reshapes Tissue-Resident Immunity in Celiac Disease

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Polylactic acid: synthesis and biomedical applications

Social and economic development has driven considerable scientific and engineering efforts on the discovery, development and utilization of polymers. Polylactic acid (PLA) is one of the most promising biopolymers as it can be produced from nontoxic renewable feedstock. PLA has emerged as an important polymeric material for biomedical applications on account of its properties such as biocompatibility, biodegradability, mechanical strength and process ability. Lactic acid (LA) can be obtained by fermentation of sugars derived from renewable resources such as corn and sugarcane. PLA is thus an eco-friendly nontoxic polymer with features that permit use in the human body. Although PLA has a wide spectrum of applications, there are certain limitations such as slow degradation rate, hydrophobicity and low impact toughness associated with its use. Blending PLA with other polymers offers convenient options to improve associated properties or to generate novel PLA polymers/blends for target applications. A variety of PLA blends have been explored for various biomedical applications such as drug delivery, implants, sutures and tissue engineering. PLA and their copolymers are becoming widely used in tissue engineering for function restoration of impaired tissues due to their excellent biocompatibility and mechanical properties. The relationship between PLA material properties, manufacturing processes and development of products with desirable characteristics is described in this article. LA production, PLA synthesis and their applications in the biomedical field are also discussed.     

Vitamin E succinate inhibits survivin and induces apoptosis in pancreatic cancer cells

Pancreatic cancer is the fourth leading cause of cancer-related deaths in the United States. Identifying novel chemotherapeutic and chemopreventive approaches is critical in the prevention and treatment of cancers such as pancreatic cancer. Vitamin E succinate (VES) is a redox-silent analog of the fat-soluble vitamin alpha-tocopherol. In the present study, we explored the antiproliferative action of VES and its effects on inhibitor of apoptosis proteins in pancreatic cancer cells. We show that VES inhibits cell proliferation and induces apoptosis in pancreatic cancer cells. Further, we demonstrate that VES downregulates the expression of survivin and X-linked inhibitor of apoptosis proteins. The apoptosis induced by VES was augmented by siRNA-mediated inhibition of survivin in PANC-1 cells. In summary, our results suggest that VES targets survivin signaling and induces apoptosis in pancreatic cancer cells.     

Serratia odorifera a Midgut Inhabitant of Aedes aegypti Mosquito Enhances Its Susceptibility to Dengue-2 Virus

Mosquito midgut plays a crucial role in its vector susceptibility and pathogen interaction. Identification of the sustainable microflora of the midgut environment can therefore help in evaluating its contribution in mosquito-pathogen interaction and in turn vector competence. To understand the bacterial diversity in the midgut of Aedes aegypti mosquitoes, we conducted a screening study of the gut microbes of these mosquitoes which were either collected from fields or reared in the laboratory "culture-dependent" approach. This work demonstrated that the microbial flora of larvae and adult Ae. aegypti midgut is complex and is dominated by Gram negative proteobacteria. Serratia odorifera was found to be stably associated in the midguts of field collected and laboratory reared larvae and adult females. The potential influence of this sustainable gut microbe on DENV-2 susceptibility of this vector was evaluated by co-feeding S. odorifera with DENV-2 to adult Ae. aegypti females (free of gut flora). The observations revealed that the viral susceptibility of these Aedes females enhanced significantly as compared to solely dengue-2 fed and another gut inhabitant, Microbacterium oxydans co-fed females. Based on the results of this study we proposed that the enhancement in the DENV-2 susceptibility of Ae. aegypti females was due to blocking of prohibitin molecule present on the midgut surface of these females by the polypeptide of gut inhabitant S. odorifera.     

Dyrk1 inhibition improves Alzheimer's disease‐like pathology

There is an urgent need for the development of new therapeutic strategies for Alzheimer's disease (AD). The dual‐specificity tyrosine phosphorylation‐regulated kinase‐1A (Dyrk1a) is a protein kinase that phosphorylates the amyloid precursor protein (APP) and tau and thus represents a link between two key proteins involved in AD pathogenesis. Furthermore, Dyrk1a is upregulated in postmortem human brains, and high levels of Dyrk1a are associated with mental retardation. Here, we sought to determine the effects of Dyrk1 inhibition on AD‐like pathology developed by 3xTg‐AD mice, a widely used animal model of AD. We dosed 10‐month‐old 3xTg‐AD and nontransgenic (NonTg) mice with a Dyrk1 inhibitor (Dyrk1‐inh) or vehicle for eight weeks. During the last three weeks of treatment, we tested the mice in a battery of behavioral tests. The brains were then analyzed for the pathological markers of AD. We found that chronic Dyrk1 inhibition reversed cognitive deficits in 3xTg‐AD mice. These effects were associated with a reduction in amyloid‐β (Aβ) and tau pathology. Mechanistically, Dyrk1 inhibition reduced APP and insoluble tau phosphorylation. The reduction in APP phosphorylation increased its turnover and decreased Aβ levels. These results suggest that targeting Dyrk1 could represent a new viable therapeutic approach for AD.     

A new mouse mutant, skijumper

Low blood sugar levels are a well-known cause of severe illness and often death in newborn humans, especially those that are small for age. Few of the causes of neonatal hypoglycemia are known, and many remain to be found. We describe a novel mouse mutant, skijumper (skimp), in which pups, despite feeding well, have low levels of glucose and develop opisthotonos, followed by death typically within a few days after birth. Genetic mapping studies have localized the lesion to a approximately 1 cM interval on mouse Chromosome (Chr) 7 between D7Mit318 and D7Mit93. We have carried out extensive analysis to define the phenotype and its likely cause. In addition to low blood glucose, affected skijumper mice have lowglycogen and ketone levels. Mass spectrometric analysis of blood samples has excluded major defects in amino acid metabolism. Initial biochemical analyses suggested a defect in ketogenesis as one possible cause of this phenotype. However, measurements of levels and activities of carnitine, carnitine palmitoyl transferases, and other enzymes involved in ketogenesis, along with studies of mitochondrial structure and function, did not demonstrate significant differences between skijumper, unaffected littermates, and control wild-type mice. These results indicate that abnormal enzyme activity in known pathways does not appear to be the primary biochemical lesion in skijumper. The skijumper may be a new valuable model for studying and understanding one type of neonatal morbidity and death.