Venom composition and pain-causing toxins of the Australian great carpenter bee Xylocopa aruana.

Most species of bee are capable of delivering a defensive sting which is often painful. A solitary lifestyle is the ancestral state of bees and most extant species are solitary, but information on bee venoms comes predominantly from studies on eusocial species. In this study we investigated the venom composition of the Australian great carpenter bee, Xylocopa aruana Ritsema, 1876. We show that the venom is relatively simple, composed mainly of one small amphipathic peptide (XYTX1-Xa1a), with lesser amounts of an apamin homologue (XYTX2-Xa2a) and a venom phospholipase-A2 (PLA2). XYTX1-Xa1a is homologous to, and shares a similar mode-of-action to melittin and the bombilitins, the major components of the venoms of the eusocial Apis mellifera (Western honeybee) and Bombus spp. (bumblebee), respectively. XYTX1-Xa1a and melittin directly activate mammalian sensory neurons and cause spontaneous pain behaviours in vivo, effects which are potentiated in the presence of venom PLA2. The apamin-like peptide XYTX2-Xa2a was a relatively weak blocker of small conductance calcium-activated potassium (KCa) channels and, like A. mellifera apamin and mast cell-degranulating peptide, did not contribute to pain behaviours in mice. While the composition and mode-of-action of the venom of X. aruana are similar to that of A. mellifera, the greater potency, on mammalian sensory neurons, of the major pain-causing component in A. mellifera venom may represent an adaptation to the distinct defensive pressures on eusocial Apidae.

Inhibit inflammation and apoptosis of pyrroloquinoline on spinal cord injury in rat.

BACKGROUND: Pyrroloquinoline quinone (PQQ) is a redox cofactor that can participate in a variety of physiological and biochemical processes, such as anti-inflammatory, cytoprotection, anti-aging, and anti-apoptosis. PQQ plays an important protective role in the central nervous system (CNS). However, the effects of PQQ on astrocytes of the CNS and spinal cord injury (SCI) of rats is still unclear. The present study investigates the role of PQQ in inflammation, apoptosis, and autophagy after SCI in rats. And the effect of PQQ on lipopolysaccharide (LPS)-induced apoptosis and inflammation of astrocytes in vitro, to explore the neuroprotective mechanism of PQQ. METHODS: Sixty specific pathogen free (SPF) SD male rats (200-250 g) were randomly divided into Normal group, Sham group, SCI group, and SCI + PQQ group, with 15 rats in each group. BBB score, HE staining, Nissl staining, Western blot, immunofluorescence, and other methods were used for detection. RESULTS: Our results showed that PQQ could upregulate BBB score in SCI rats. In the second place, PQQ can increase the number and improve the morphology of neurons after SCI. The expression of IL-1ß, TNF-α, IL-6 was significantly decreased after PQQ treatment. And then, the ratio of B-cell lymphoma-2 (Bcl-2)/Bcl-2 associated X protein (Bax) increased significantly, and the positive signal of NeuN increased obviously after PQQ treatment. There are a large number of co-localizations between Bcl-2 and NeuN. Meanwhile, PQQ could down-regulate the expression of Active-Caspase3, and PQQ treatment could reverse the transfer of Active-Caspase3/Caspase3 from the cytoplasm to the nucleus in neurons and astrocytes after SCI. At the same time, PQQ had no significant effect on the LC3b/a ratio. PQQ could decrease the LAMP2 expression in spinal cord after injury. The expression level of phospho-Akt (p-AKT) increased after SCI and decreased after PQQ treatment. In primary astrocytes, LPS could induce the expression levels of IL-1ß, TNF-α, and IL-6, and which were inhibited by PQQ treatment at 12 hours. After treatment with LPS, the expression level of Active-Caspase3 increased, which could be reversed by PQQ treatment for 24 h. CONCLUSIONS: These results suggest that PQQ can ameliorate the motor function of hind limbs and the pathological changes of neurons and injured spinal cord after SCI, down-regulate the expressions of IL-1ß, TNF-α, and IL-6, inhibit apoptosis after SCI, and inhibit LPS-induced apoptosis and inflammation of astrocytes.

Effect of pyrroloquinoline quinone on lipopolysaccharide-induced autophagy in HAPI microglia cells.

BACKGROUND: Pyrroloquinoline quinone (PQQ) is involved in various physiological and biochemical processes, including antioxidant, cell proliferation, and mitochondrial formation. It plays a vital role in protecting neurons. However, the effect of PQQ on microglia, an inflammatory cell of the central nervous system (CNS), is still unclear. This study aimed to investigate the biological role and neuroprotective mechanism of PQQ in HAPI microglial cells exposed to lipopolysaccharide (LPS). METHODS: Western blot (WB) was used to detect apoptosis and autophagy-related molecules Bax, Bcl2, active-caspase-3, caspase-3, LC3, lysosomal associated membrane protein 2 (LAMP2), AKT, tumor necrosis factor receptor (TNFR) 1, and TNFR2 expression. The phosphatidylinositol 3-kinase (PI3K)/Akt inhibitor LY294002 was used to block the Akt pathway. WB detected the effects of PI3K on autophagy and TNFR1 and TNFR2 expression. The localization of active-caspase-3, caspase-3, LC3, LAMP2, TNFR1, and TNFR2 in cells was observed by immunofluorescence staining. The effect of PQQ on the cell cycle was examined by flow cytometry. We used 5-Ethynyl-2'-deoxyuridine (EdU) assay to detect cell proliferation. The migration ability of cells under different conditions was detected by scratch test and Transwell assay. RESULTS: Our results showed that there were different effects on the apoptosis-related molecules Bcl2/Bax and active-caspase-3/caspase in HAPI microglial cells treated with PQQ at different times. PQQ had no significant effect on the LC3b/a ratio in the early stage, which was upregulated in the later stage. The expression of LAMP2 was significantly increased in both early and late stages after PQQ treatment. At the same time, we found that PQQ can reverse the translocation of LAMP2 from the cytoplasm to the nucleus in LPS-induced HAPI microglia. After PQQ treatment, TNFR1 was significantly decreased, but TNFR2 increased in LPS-induced HAPI microglia. It may be that PQQ works through the PI3K/Akt signaling pathway to up-regulate LC3, LAMP2, and TNFR1 and down-regulate TNFR2 in LPS-induced HAPI microglia. However, PQQ has little effect on LPS-induced proliferation, cell cycle, and migration of HAPI microglia. CONCLUSIONS: In LPS-induced HAPI microglia, PQQ reduces the apoptosis level and increases that of autophagy. In addition, PQQ changes the distribution of LAMP2 in the cytoplasm and nucleus, which is regulated through the PI3K/Akt signaling pathway.

The Impact of ß-1,4-Galactosyltransferase V on Microglial Function.

ß-1,4 Galactosyltransferase V (ß-1,4-GalT V) belongs to the ß-1,4 galactosyltransferase family, which modifies proteins and plays a vital role in biological function. Our previous study revealed that ß-1,4-GalT V was expressed in the cortex and hippocampus and participated in the recovery of spatial learning and memory in rats with traumatic brain injury. However, the expression of ß-1,4-GalT V in microglia, resident immune cells in the central nervous system, and its impact on microglia in resting and lipopolysaccharide-triggered activated stages are elusive. In this study, we clarified that ß-1,4-GalT V expresses in microglia, and it regulates microglial migration, proliferation, and release of the inflammatory factors. We also observed that ß-1,4-GalT V affects the expression level of tumor necrosis factor receptor (TNFR)2 instead of TNFR1. These results strongly support the fact that ß-1,4-GalT V is involved in microglial function.