Myocardial SDF-1 expression was increased only in the early phase post MI. MSCs intravenous infused at the early phase of MI were recruited to injured heart, enhanced angiogenesis and improved cardiac function.

Detergents have been widely used for the solubilization of membrane proteins and the improvement of their digestion. In this paper, we have evaluated the application of sodium deoxycholate (SDC) to the solubilization and digestion of rat hippocampal plasma membrane (PM) proteins. For in-solution digestion, rat hippocampal PM fraction from sucrose-density gradient centrifugation was solubilized by boiling in 1.0% SDC, and directly digested without dilution. During the in-gel digestion of the hippocampal PM proteins separated by SDS-PAGE, 0.1% SDC was added. Before analysis of peptide mixture by liquid chromatography and electrospray mass spectrometry, SDC in the tryptic digests was removed by centrifugation following acidification. Use of 1.0% SDC in solubilization and in-solution digestion of rat PM proteins had led to 77 PM or membrane-associated proteins identified, a more than 2-fold increase over that by use of SDS. The addition of 0.1% SDC to the in-gel digestion of SDS-PAGE-resolved membrane proteins remarkably enhanced the coverage of tryptic peptides and the number of hydrophobic membrane proteins identified. Being a cheaper and more tractable acid-insoluble detergent, SDC could be used at higher concentration in the solubilization and tryptic digestion of proteins including PM proteins with the purpose of enhancing the protein solubility and at the same time making no interference with trypsin activity and subsequent analyses.

When developing malaria vaccines, the most crucial step is to elucidate the mechanisms involved in protective immunity against the parasites. We found that CD8+ T cells contribute to protective immunity against infection with blood‐stage parasites of Plasmodium yoelii. Infection of C57BL/6 mice with P. yoelii 17XL was lethal, while all mice infected with a low‐virulence strain of the parasite 17XNL acquired complete resistance against re‐infection with P. yoelii 17XL. However, the host mice transferred with CD8+ T cells from mice primed only with P. yoelii 17XNL failed to acquire protective immunity. On the other hand, the irradiated host mice were completely resistant to P. yoelii 17XL infection, showing no grade of parasitemia when adoptively transferred with CD8+ T cells from immune mice that survived infection with both P. yoelii XNL and, subsequently, P. yoelii 17XL. These protective CD8+ T cells from immune WT mice had the potential to generate IFN‐γ, perforin (PFN) and granzyme B. When mice deficient in IFN‐γ were used as donor mice for CD8+ T cells, protective immunity in the host mice was fully abrogated, and the immunity was profoundly attenuated in PFN‐deficient mice. Thus, CD8+ T cells producing IFN‐γ and PFN appear to be involved in protective immunity against infection with blood‐stage malaria.

Malaria is still a life-threatening infectious disease that continues to produce 2 million deaths annually. Malaria parasites have acquired immune escape mechanisms and prevent the development of sterile immunity. Regulatory T cells (Tregs) have been reported to contribute to immune evasion during malaria in mice and humans, suggesting that activating Tregs is one of the mechanisms by which malaria parasites subvert host immune systems. However, little is known about how these parasites activate Tregs. We herein show that TLR9 signaling to dendritic cells (DCs) is crucial for activation of Tregs. Infection of mice with the rodent malaria parasite Plasmodium yoelii activates Tregs, leading to enhancement of their suppressive function. In vitro activation of Tregs requires the interaction of DCs with parasites in a TLR9-dependent manner. Furthermore, TLR9−/− mice are partially resistant to lethal infection, and this is associated with impaired activation of Tregs and subsequent development of effector T cells. Thus, malaria parasites require TLR9 to activate Tregs for immune escape.

SummaryAll 21 of the Nudix hydrolase genes from the radiation-resistant organism Deinococcus radiodurans have been cloned into vectors under the control of T7 promoters and expressed as soluble proteins in Escherichia coli. Their sizes range from 9.8 kDa (91 amino acids) to 59 kDa (548 amino acids). Two novel proteins were identified, each with two Nudix boxes in its primary structure, unique among all other known Nudix hydrolases. Extracts of each of the expressed proteins were assayed by a generalized procedure that measures the hydrolysis of nucleoside diphosphate derivatives, and several enzymatic activities were tentatively identified. In addition to representatives of known Nudix hydrolase subfamilies active on ADP-ribose, NADH, dinucleoside polyphosphates or (deoxy)nucleoside triphosphates, two new enzymes, a UDP-glucose pyrophosphatase and a CoA pyrophosphatase, were identified.

Delta (Y), MB1 (X), and Z are the three catalytic beta-subunits located in the inner rings of the constitutive proteasome, an intracellular multicatalytic complex responsible for the generation of peptides presented by human leukocyte antigen (HLA) class I antigens to T cells. When cells are incubated with interferon-gamma, delta (Y), MB1 (X), and Z are replaced by LMP2, LMP7, and LMP10, respectively, leading to the expression of immunoproteasome which generates peptides with increased affinity for HLA class I antigens. The characterization of the expression of constitutive proteasome and immunoproteasome subunits in cells, normal tissues, and malignant lesions has been hampered by the lack or limited availability of constitutive proteasome and immunoproteasome subunit-specific monoclonal antibodies (mAbs), which are suitable for immunohistochemical staining. To overcome this limitation, we generated human delta (Y), MB1 (X), Z, LMP2, LMP7, and LMP10-specific mAb-secreting hybridomas from BALB/c mice immunized with peptides and recombinant fusion proteins. The mAbs SY-5, SJJ-3, NB-1, SY-1, HB-2, and TO-7 were shown to be specific for delta (Y), MB1 (X), Z, LMP2, LMP7, and LMP10, respectively, as they react specifically with the corresponding molecules when tested with a human B lymphoid LG2 cell lysate in Western blotting and with the peptide derived from each molecule in enzyme-linked immunosorbent assay. The reactivity of the six mAbs with the corresponding intracellular antigens resulted in intracellular staining when the mAbs were tested with microwave-treated and saponin-permeabilized cells in indirect immunofluorescence and with formalin-fixed, paraffin-embedded tissue sections in immunohistochemical reactions. These results suggest that the constitutive proteasome and immunoproteasome subunit-specific mAbs we have developed are useful probes to characterize the expression of proteasome subunits in normal tissues and in pathological lesions.

N-acetylserotonin (NAS) is synthesized from serotonin by arylalkylamine N-acetyltransferase (AANAT), which is predominantly expressed in the pineal gland and retina. NAS activates TrkB in a circadian manner and exhibits antidepressant effects in a TrkB-dependent manner. It also enhances neurogenesis in hippocampus in sleep-deprived mice. Here we report the identification of NAS derivatives that possess much more robust neurotrophic effects with improved pharmacokinetic profiles. The compound N-[2-(5-hydroxy-1H-indol-3-yl)ethyl]-2-oxopiperidine-3-carboxamide (HIOC) selectively activates TrkB receptor with greater potency than NAS. It potently protects retinas from light-induced retinal degeneration (LIRD), which is tightly coupled with pronounced TrkB activation in retinas. Pharmacokinetic studies demonstrate that this compound is stable in serum and liver microsomes. It can pass the blood-brain barrier and bloodretinal barrier. Hence, HIOC is a good lead compound for further drug development for treating retinal degenerative diseases.circadian rhythm | melatonin | small molecule | neurotrophins A rylalkylamine N-acetyltransferase (AANAT) acetylates Nacetylserotonin (NAS) from the neurotransmitter serotonin in the pineal gland and retina. NAS is subsequently methylated and converted into melatonin by hydroxyindole-O-methyltransferase (HIOMT). The three components of the melatonin synthetic pathway, namely 5-HT, NAS, and melatonin, all display dramatic circadian rhythms (1). It has long been thought that NAS only acts as a precursor of melatonin in the process of melatonin biosynthesis. Recently, we demonstrated that NAS activates the TrkB receptor and exerts antidepressant effects in a TrkB-dependent manner (2). Moreover, we reported that NAS but not melatonin stimulates neurogenesis in the hippocampus of sleep-deprived mice via activating TrkB receptor in dentate gyrus (3). In addition to these findings, NAS has been shown to facilitate memory (4), regulate hypothermic body temperature (5), induce analgesia in central nervous system (6), and exert antioxidative actions (7).Brain-derived neurotrophic factor (BDNF), a member of the neurotrophin family that also includes nerve growth factor (NGF), Neurotrophin-3 (NT-3), and NT-4/5, exerts its biological functions through two transmembrane receptors: the p75 neurotrophin receptor (p75NTR) and TrkB receptor tyrosine kinase. BDNF binding to TrkB triggers its dimerization and autophosphorylation of tyrosine residues in its intracellular domain, leading to activation of the three major downstream signaling cascades, including mitogen-activated protein kinase (MAPK), phosphatidylinositol 3-kinase (PI3K), and phospholipase C-γ1 (PLC-γ1) (8, 9). Through these pathways, BDNF mediates a variety of neuronal activities involved in neuronal survival, neurogenesis, and synaptic plasticity. Accumulating evidence demonstrates that neurotrophins play critical roles in the development of the retina and visual system (10-12). Limiting amounts of neurotrophins in the target area of th...

a b s t r a c tThe tropomyosin-related kinase A (TrkA) receptor and its ligand, nerve growth factor (NGF), play crucial roles in the development and function of the nervous system. NGF is believed to activate TrkA by bridging two TrkA monomers, leading to TrkA transphosphorylation. However, here we show that the majority of TrkA receptors exist as preformed, yet inactive, homodimers prior to NGF binding by using three different approaches such as chemical crosslinking and enzyme fragment complementation assay. Furthermore, TrkA homodimers are formed in endoplasmic reticulum before newly synthesized receptors reach the cell surface. These findings shed light on molecular mechanisms underlying transmembrane signaling by TrkA. Structured summary:TrkA physically interacts with TrkA by protein complementation assay (View interaction) TrkA physically interacts with TrkA by bimolecular fluorescence complementation (View interaction) TrkA physically interacts with TrkA by cross-linking study (View interaction)