Platelet activating factor induces transient blood‐brain barrier opening to facilitate edaravone penetration into the brain

The blood–brain barrier (BBB) greatly limits the efficacy of many neuroprotective drugs' delivery to the brain, so improving drug penetration through the BBB has been an important focus of research. Here we report that platelet activating factor (PAF) transiently opened BBB and facilitated neuroprotectant edaravone penetration into the brain. Intravenous infusion with PAF induced a transient BBB opening in rats, reflected by increased Evans blue leakage and mild edema formation, which ceased within 6 h. Furthermore, rat regional cerebral blood flow (rCBF) declined acutely during PAF infusion, but recovered slowly. More importantly, this transient BBB opening significantly increased the penetration of edaravone into the brain, evidenced by increased edaravone concentrations in tissue interstitial fluid collected by microdialysis and analyzed by Ultra‐performance liquid chromatograph combined with a hybrid quadrupole time‐of‐flight mass spectrometer (UPLC‐MS/MS). Similarly, incubation of rat brain microvessel endothelial cells monolayer with 1 μM PAF for 1 h significantly increased monolayer permeability to ¹²⁵I‐albumin, which recovered 1 h after PAF elimination. However, PAF incubation with rat brain microvessel endothelial cells for 1 h did not cause detectable cytotoxicity, and did not regulate intercellular adhesion molecule‐1, matrix‐metalloproteinase‐9 and P‐glycoprotein expression. In conclusion, PAF could induce transient and reversible BBB opening through abrupt rCBF decline, which significantly improved edaravone penetration into the brain.

     

Early secreted antigenic target of 6-kDa of Mycobacterium tuberculosis promotes caspase-9/caspase-3-mediated apoptosis in macrophages

Molecular and Cellular Biochemistry - Mycoplasma pneumoniae pneumonia (MPP) is the most common respiratory infection in young children and its incidence has increased worldwide. In this study, high...     

A survey of the methods for the characterization of microbial consortia and communities

A survey of the available literature on methods most frequently used for the identification and characterization of microbial strains, communities, or consortia is presented. The advantages and disadvantages of the various methodologies were examined from several perspectives including technical, economic (time and cost), and regulatory. The methods fall into 3 broad categories: molecular biological, biochemical, and microbiological. Molecular biological methods comprise a broad range of techniques that are based on the analysis and differentiation of microbial DNA. This class of methods possesses several distinct advantages. Unlike most other commonly used methods, which require the production of secondary materials via the manipulation of microbial growth, molecular biological methods recover and test their source materials (DNA) directly from the microbial cells themselves, without the requirement for culturing. This eliminates both the time required for growth and the biases associated with cultured growth, which is unavoidably and artificially selective. The recovered nucleic acid can be cloned and sequenced directly or subpopulations can be specifically amplified using polymerase chain reaction (PCR), and subsequently cloned and sequenced. PCR technology, used extensively in forensic science, provides researchers with the unique ability to detect nucleic acids (DNA and RNA) in minute amounts, by amplifying a single target molecule by more than a million-fold. Molecular methods are highly sensitive and allow for a high degree of specificity, which, coupled with the ability to separate similar but distinct DNA molecules, means that a great deal of information can be gleaned from even very complex microbial communities. Biochemical methods are composed of a more varied set of methodologies. These techniques share a reliance on gas chromatography and mass spectrometry to separate and precisely identify a range of biomolecules, or else investigate biochemical properties of key cellular biomolecules. Like the molecular biological methods, some biochemical methods such as lipid analyses are also independent of cultured growth. However, many of these techniques are only capable of producing a profile that is characteristic of the microbial community as a whole, providing no information about individual members of the community. A subset of these methodologies are used to derive taxonomic information from a community sample; these rely on the identification of key subspecies of biomolecules that differ slightly but characteristically between species, genera, and higher biological groupings. However, when the consortium is already growing in chemically defined media (as is often the case with commercial products), the rapidity and relatively low costs of these procedures can mitigate concerns related to culturing biases. Microbiological methods are the most varied and the least useful for characterizing microbial consortia. These methods rely on traditional tools (cell counting, selective growth, and microscopic examination) to provide more general characteristics of the community as a whole, or else to narrow down and identify only a small subset of the members of that community. As with many of the biochemical methods, some of the microbiological methods can fairly rapidly and inexpensively create a community profile, which can be used to compare 2 or more entire consortia. However, for taxonomic identification of individual members, microbiological methods are useful only to screen for the presence of a few key predetermined species, whose preferred growth conditions and morphological characteristics are well defined and reproducible.     

Arylphthalazines: identification of a new phthalazine chemotype as inhibitors of VEGFR kinase

A novel class of 4-arylamino-phthalazin-1-yl-benzamides is described as inhibitors of vascular endothelial growth factor receptor II (VEGFR-2). Several compounds display potent VEGFR-2 inhibitory activity with an IC50 as low as 0.078 microM in an HTRF enzymatic assay. These compounds are relatively selective against a small kinase panel.     

Prevention of meningo/encephalomyelitis due to Sarcocystis neurona infection in mice is mediated by CD8 cells

Immunodeficient CD8 knockout mice were infected with Sarcocystis neurona merozoites, in order to determine the role of CD8 cells in protective immunity. Using a direct agglutination test, all infected mice seroconverted by selected time points. Infected mice developed splenomegaly and bilateral lymphadenopathy. Histological changes included marked follicular development in the spleen, endothelitis and moderate perivascular inflammation in the liver, and meningoencephalitis in the brain. Infected brains were positive for S. neurona by polymerase chain reaction. Corresponding to histopathological changes, there were decreased numbers of B-cells in the spleen. The mice did not have significant memory (CD44hi/CD4) or effector (CD45RBhi/CD4) populations present at the time of euthanasia. Flow cytometry confirmed the lack of CD8 cells. Taken together, these data support previous studies suggesting a critical role for CD8 cells in the prevention of menigoencephalitis in S. neurona-infected mice.     

Specific cleavage of Mcl-1 by caspase-3 in tumor necrosis factor-related apoptosis-inducing ligand (TRAIL)-induced apoptosis in Jurkat leukemia T cells

Tumor necrosis factor-related apoptosis-inducing ligand (TRAIL) induces programmed cell death through the caspase activation cascade and translocation of cleaved Bid (tBid) by the apical caspase-8 to mitochondria to induce oligomerization of multidomain Bax and Bak. However, the roles of prosurvival Bcl-2 family proteins in TRAIL apoptosis remain elusive. Here we showed that, besides the specific cleavage and activation of Bid by caspase-8 and caspase-3, TRAIL-induced apoptosis in Jurkat T cells required the specific cleavage of Mcl-1 at Asp-127 and Asp-157 by caspase-3, while other prototypic antiapoptotic factors such as Bcl-2 or Bcl-X(L) seemed not to be affected. Mutation at Asp-127 and Asp-157 of Mcl-1 led to cellular resistance to TRAIL-induced apoptosis. In sharp contrast to cycloheximide-induced Mcl-1 dilapidation, TRAIL did not activate proteasomal degradation of Mcl-1 in Jurkat cells. We further established for the first time that the C-terminal domain of Mcl-1 became proapoptotic as a result of caspase-3 cleavage, and its physical interaction and cooperation with tBid, Bak, and voltage-dependent anion-selective channel 1 promoted mitochondrial apoptosis. These results suggested that removal of N-terminal domains of Bid by caspase-8 and Mcl-1 by caspase-3 enabled the maximal mitochondrial perturbation that potentiated TRAIL-induced apoptosis.     

Immunogenicity of outer membrane derivatives ofHaemophilus influenzae type b

We prepared outer membrane derivatives ofHaemophilus influenzae type b to determine whether the residual capsular and noncapsular surface components are immunogenic and protective. These fragments consist primarily of six major proteins and lipopolysaccharide. By transmission electron microscopy, they appeared as small, membrane-like fragments or larger, cellshaped double-track ghosts. Rabbits immunized with ghosts responded with increases in serum anticapsular antibody and bactericidal activity. Antisera absorbed with capsular antigen to remove anticapsular antibody remained bactericidal and passively protected infant rats. These data suggest that antibodies to noncapsular surface antigens are protective, and that outer membrane derivatives retain some of the constituents responsible for stimulating immunity.