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Background: The descending noradrenergic (NAergic) system is one of the important endogenous analgesia systems. It has been suggested that noxious stimuli could activate descending NAergic system; nevertheless, the underlying neuronal circuit remains unclear. As NAergic neurons in the A7 catecholamine cell group (A7) are a part of the descending NAergic system and the lateral parabrachial nucleus (LPB) is an important brainstem structure that relays ascending nociceptive signal, we aimed to test whether LPB neurons have direct synaptic contact with NAergic A7 neurons., Results: Stereotaxic injections of an anterograde tracer, biotinylated dextran-amine (BDA), were administered to LPB in rats. The BDA-labeled axonal terminals that have physical contacts with tyrosine hydroxylase-positive (presumed noadrenergic) neurons were identified in A7. Consistent with these morphological observations, the excitatory synaptic currents (EPSCs) were readily evoked in NAergic A7 neurons by extracellular stimulation of LPB. The EPSCs evoked by LPB stimulation were blocked by CNQX, a non-NMDA receptor blocker, and AP5, a selective NMDA receptor blocker, showing that LPB-A7 synaptic transmission is glutamatergic. Moreover, the amplitude of LPB-A7 EPSCs was significantly attenuated by DAMGO, a selective μ-opioid receptor agonist, which was associated with an increase in paired-pulse ratio., Conclusions: Taken together, the above results showed direct synaptic connections between LPB and A7 catecholamine cell group, the function of which is subject to presynaptic modulation by μ-opioid receptors.

Introduction: Idiopathic pulmonary fibrosis is a progressive diffuse parenchymal lung disorder of unknown etiology. Mesenchymal stem cell (MSC)-based therapy is a novel approach with great therapeutic potential for the treatment of lung diseases. Despite demonstration of MSC grafting, the populations of engrafted MSCs have been shown to decrease dramatically 24 hours post-transplantation due to exposure to harsh microenvironments. Hypoxia is known to induce expression of cytoprotective genes and also secretion of anti-inflammatory, anti-apoptotic and anti-fibrotic factors. Hypoxic preconditioning is thought to enhance the therapeutic potency and duration of survival of engrafted MSCs. In this work, we aimed to prolong the duration of survival of engrafted MSCs and to enhance the effectiveness of idiopathic pulmonary fibrosis transplantation therapy by the use of hypoxia-preconditioned MSCs., Methods: Hypoxic preconditioning was achieved in MSCs under an optimal hypoxic environment. The expression levels of cytoprotective factors and their biological effects on damaged alveolar epithelial cells or transforming growth factor-beta 1-treated fibroblast cells were studied in co-culture experiments in vitro. Furthermore, hypoxia-preconditioned MSCs (HP-MSCs) were intratracheally instilled into bleomycin-induced pulmonary fibrosis mice at day 3, and lung functions, cellular, molecular and pathological changes were assessed at 7 and 21 days after bleomycin administration., Results: The expression of genes for pro-survival, anti-apoptotic, anti-oxidant and growth factors was upregulated in MSCs under hypoxic conditions. In transforming growth factor-beta 1-treated MRC-5 fibroblast cells, hypoxia-preconditioned MSCs attenuated extracellular matrix production through paracrine effects. The pulmonary respiratory functions significantly improved for up to 18 days of hypoxia-preconditioned MSC treatment. Expression of inflammatory factors and fibrotic factor were all downregulated in the lung tissues of the hypoxia-preconditioned MSC-treated mice. Histopathologic examination observed a significant amelioration of the lung fibrosis. Several LacZ-labeled MSCs were observed within the lungs in the hypoxia-preconditioned MSC treatment groups at day 21, but no signals were detected in the normoxic MSC group. Our data further demonstrated that upregulation of hepatocyte growth factor possibly played an important role in mediating the therapeutic effects of transplanted hypoxia-preconditioned MSCs., Conclusion: Transplantation of hypoxia-preconditioned MSCs exerted better therapeutic effects in bleomycin-induced pulmonary fibrotic mice and enhanced the survival rate of engrafted MSCs, partially due to the upregulation of hepatocyte growth factor.

Background: Klebsiella pneumoniae displaying the hypermucoviscosity (HV) phenotype are considered more virulent than HV-negative strains. Nevertheless, the emergence of tissue-abscesses-associated HV-negative isolates motivated us to re-evaluate the role of HV-phenotype., Results: Instead of genetically manipulating the HV-phenotype of K. pneumoniae, we selected two clinically isolated K1 strains, 1112 (HV-positive) and 1084 (HV-negative), to avoid possible interference from defects in the capsule. These well-encapsulated strains with similar genetic backgrounds were used for comparative analysis of bacterial virulence in a pneumoniae or a liver abscess model generated in either naïve or diabetic mice. In the pneumonia model, the HV-positive strain 1112 proliferated to higher loads in the lungs and blood of naïve mice, but was less prone to disseminate into the blood of diabetic mice compared to the HV-negative strain 1084. In the liver abscess model, 1084 was as potent as 1112 in inducing liver abscesses in both the naïve and diabetic mice. The 1084-infected diabetic mice were more inclined to develop bacteremia and had a higher mortality rate than those infected by 1112. A mini-Tn5 mutant of 1112, isolated due to its loss of HV-phenotype, was avirulent to mice., Conclusion: These results indicate that the HV-phenotype is required for the virulence of the clinically isolated HV-positive strain 1112. The superior ability of the HV-negative stain 1084 over 1112 to cause bacteremia in diabetic mice suggests that factors other than the HV phenotype were required for the systemic dissemination of K. pneumoniae in an immunocompromised setting.

Background: Detailed knowledge of the molecular and cellular mechanisms that direct spatial and temporal gene expression in pre-implantation embryos is critical for understanding the control of the maternal-zygotic transition and cell differentiation in early embryonic development. In this study, twenty-three clones, expressed at different stages of early mouse development, were identified using differential display reverse transcription polymerase chain reaction (DDRT-PCR). One of these clones, which is expressed in 2-cell stage embryos at 48 hr post-hCG injection, shows a perfect sequence homology to the gene encoding the granzyme G protein. The granzyme family members are serine proteases that are present in the secretory granules of cytolytic T lymphocytes. However, the pattern of granzyme G expression and its function in early mouse embryos are entirely unknown., Results: Upon the introduction of an antisense morpholino (2 mM) against granzyme G to knock-down endogenous gene function, all embryos were arrested at the 2- to 4-cell stages of egg cleavage, and the de novo synthesis of zygotic RNAs was decreased. The embryonic survival rate was dramatically decreased at the late 2-cell stage when serine protease-specific inhibitors, 0.1 mM 3,4-dichloroisocoumarin (3,4-DCI), and 2 mM phenyl methanesulphonyl fluoride (PMSF), were added to the in vitro embryonic culture medium. Survival was not affected by the addition of 0.5 mM EDTA, a metalloproteinase inhibitor., Conclusion: We characterized for the first time the expression and function of granzyme G during early stage embryogenesis. Our data suggest that granzyme G is an important factor in early mouse embryonic development and may play a novel role in the elimination of maternal proteins and the triggering of zygotic gene expression during the maternal-zygotic transition.