Supraspinal antinociceptive effect of apelin-13 in a mouse visceral pain model

Apelin, as the endogenous ligand of the APJ receptor, is a novel identified neuropeptide whose biological functions are not fully understood. APJ receptor mRNA was found in several brain regions related to descending control system of pain, such as amygdala, hypothalamus and dorsal raphe nucleus (DRN). The present study was designed to determine whether supraspinal apelin-13 may produce antinociceptive effect observed in the acetic acid-induced writhing test, a model of visceral pain. Apelin-13 not only significantly produced preemptive antinociception at the dose of 0.3, 0.5, 1 and 3μg/mouse when injected intracerebroventricularly (i.c.v.) before acetic acid, but also significantly induced antinociception at a dose of 0.5, 1 and 3μg/mouse when injected i.c.v. after acetic acid. And i.c.v. apelin-13 did not influence 30-min locomotor activity counts in mice. Intrathecal (i.t.) administration of apelin-13 (1 and 3μg/mouse) significantly decreased the number of writhes, however, intraperitoneal (i.p.) injection of apelin-13 (10-100μg/mouse) had no effect on the number of writhes in the writhing test. The specific APJ receptor antagonist apelin-13(F13A), no-specific opioid receptor antagonist naloxone and μ-opioid receptor antagonist β-funaltrexamine hydrochloride (β-FNA) could significantly antagonize the antinociceptive effect of i.c.v. apelin-13, suggesting APJ receptor and μ-opioid receptor are involved in this process. Central low dose of apelin-13 (0.3μg/mouse, i.c.v.) could significantly potentiate the analgesic potencies of modest and even relatively ineffective doses of morphine administrated at supraspinal level. This enhanced antinociceptive effect was reversed by naloxone, suggesting that the potentiated analgesic response is mediated by opioid-responsive neurons.     

Nemo-like Kinase (NLK) Involves in Neuronal Apoptosis after Traumatic Brain Injury

Traumatic brain injury (TBI) consists of two phases: an immediate phase in which damage is caused as a direct result of the mechanical impact: and a late phase of altered biochemical events that results in delayed tissue damage and is therefore amenable to therapeutic treatment. Because the molecular mechanisms of delayed post-traumatic neuronal cell death are still poorly understood, we investigated whether nemo-like kinase (NLK), an evolutionarily conserved serine/threonine kinase involved in neuronal apoptosis following TBI. In the model of TBI, western blot analysis, double immunofluorescent staining and immunohistochemistry were used to analyze the role of NLK in the process. The results showed a significant down-regulation of NLK and a concomitant up-regulation of caspase-3 during the early stage of TBI. In the model of glutamate inducing PC12 apoptosis, we analyzed the effect of over-expression of NLK on the neuronal cell line PC12 apoptosis by cck-8, western blot and TUNEL assays. Together with previous reports. We hypothesize NLK was related to the down-regulation of caspase-3 expression after TBI, and such an event may be associated with neuronal apoptosis.     

A novel strategy of natural plant ferritin to protect DNA from oxidative damage during iron oxidation

Plant ferritin is a naturally occurring heteropolymer in plastids, where Fe(2+) is oxidatively deposited into the protein. However, the effect of this process on the coexistence of DNA and plant ferritin in the plastids is unknown. To investigate this effect, we built a system in which various plant ferritins and DNA coexist, followed by treatment with ferrous ions under aerobic conditions. Interestingly, naturally occurring soybean seed ferritin (SSF), a heteropolymer with an H-1/H-2 ratio of 1 to 1 in the apo form, completely protected DNA from oxidative damage during iron oxidative deposition into protein, and a similar result was obtained with its recombinant form, but not with its homopolymeric counterparts, apo rH-1 and apo rH-2. We demonstrate that the difference in DNA protection between heteropolymeric and homopolymeric plant ferritins stems from their different strategies to control iron chemistry during the above oxidative process. For example, the detoxification reaction occurs only in the presence of apo heteropolymeric SSF (hSSF), thereby preventing the production of hydroxyl radicals. In contrast, hydroxyl radicals are apparently generated via the Fenton reaction when apo rH-1 or rH-2 is used instead of apo hSSF. Thus, a combination of H-1 and H-2 subunits in hSSF seems to impart a unique DNA-protective function to the protein, which was previously unrecognized. This new finding advances our understanding of the structure and function of ferritin and of the widespread occurrence of heteropolymeric plant ferritin in nature.     

Effects of Ginkgo biloba extract on inflammatory mediators (SOD, MDA, TNF-alpha, NF-kappaBp65, IL-6) in TNBS-induced colitis in rats

Inflammatory mediators play a critical role in ulcerative colitis immune and inflammatory processes. The aim of the study was to investigate the effects of Ginkgo biloba extract on inflammatory mediators (SOD, MDA, TNF-alpha, NF-kappaBp65, IL-6) in TNBS-induced colitis in rats. Colitis in rats was induced by colonic administration with 2,4,6-trinitrobenzene sulfonic acid (TNBS, 150 mg/kg). EGB in doses of (50, 100, 200 mg/kg) was administered for 4 weeks to protect colitis. The results showed that EGB could significantly ameliorate macroscopic and histological damage, evidently elevate the activities of SOD and reduce the contents of MDA, inhibit the protein and mRNA expressions of TNF-alpha, NF-kappaBp65, and IL-6 in the colon tissues of experimental colitis in a dose-dependent manner compared with the model group. We concluded that the probable mechanisms of EGB ameliorated inflammatory injury in TNBS-induced colitis in rats by its modulation of inflammatory mediators and antioxidation.     

Abundance and diversity of aerobic anoxygenic phototrophic bacteria in saline lakes on the Tibetan plateau

Aerobic anoxygenic phototrophic (AAP) bacteria are heterotrophic prokaryotes that are capable of utilizing light as an energy source but are not capable of producing molecular oxygen. Recently, multiple studies have found that AAP bacteria are widely distributed in oceans and estuaries and may play an important role in carbon cycling. However, AAP bacteria in inland lake ecosystems have not been investigated in depth. In this study, the abundance and diversity of the pufL-M genes, encoding photosynthetic reaction centers of AAP bacteria, were determined in the oxic water column and anoxic sediments of saline lakes (Qinghai, Erhai, and Gahai Lakes) on the Tibetan Plateau, China. Our results indicated that AAP bacteria were abundant in inland lakes, with the proportion of AAP bacteria (in total bacteria) comparable to those in the oceans, but with a lower diversity. Salinity and pH were found to be potential factors controlling the AAP bacterial diversity and community composition. Our data have implications for a better understanding of the potential role of AAP bacteria in carbon cycling in inland lake ecosystems.     

A hydrophobicity-dependent motif responsible for surface expression of cardiac potassium channel

The long-QT syndrome (LQTS) is an inherited cardiac disorder associated with syncope and a high risk of sudden death. The molecular basis of type-1 LQTS (LQT1) is a missense or nonsense mutation in KCNQ channels that reduces slowly activating delayed rectifier potassium channel (I_Ks) resulting in a prolonged action potential. Noticeably, the S2–S3 linker is a highly congregating region of LQT1 mutations. To further explore the mechanism, a KCNQ mutant (L191P) identified in one Chinese pedigree with LQT1 was chosen for this purpose. As Leu-191 is located in the middle of a well-known endoplasmic reticulum (ER) localization signal (RXR) in the intracellular S2–S3 linker, we examined the kinetics and the surface expression of both the KCNQ1 and L191 mutants. Our results showed that the mutation did not affect the channel kinetics, whereas the surface expression increased with increasing hydrophobicity of the middle residue ‘X’ of the RXR motif. Based on an analysis of fractional fluorescence data using a binomial model, we also found that the percentage of KCNQ1/L191P heteromeric channels expressed at the cell surface were 22.0%, 40.5%, 27.9%, 8.6% and 1.0% of heteromeric channels with 0, 1, 2, 3 and 4 subunits of L191P, respectively, in a transfected ratio of KCNQ1: L191P = 1:1. These experiments demonstrated that coexpression of L191P resulted in a trafficking factor α < 1, causing a trafficking deficiency of heteromeric channels that underlay the dominant-negative effect. This study suggests several trafficking signals coexisting in this region, and expands our understanding of possible dominant-negative mechanisms underlying LQTS.     

Kinetic determination of tryptophan by using the B-Z oscillating chemical system

A simple and rapid method was devised for determination of tryptophan, based on the Belousov-Zhabotinskii (B-Z) oscillating chemical system. Changes in oscillating period and amplitude were linearly proportional to the negative logarithm of l-tryptophan concentration over the range of 6.44 × 10⁻⁷–2.55 × 10⁻⁴ M, with the regression coefficients of near unity and a lower detection limit of 6.5 × 10⁻⁸ M. d-tryptophan was also examined although it is rarely found in most biological fluids, and perhaps not at all in natural proteins. The change of period against to negative logarithm of d-tryptophan concentration over the range of 4.9 × 10⁻⁵–8.24 × 10⁻⁴ M is linear. Because the optimum conditions for determination of l- and d-tryptophan are not the same, a little amount of d-tryptophan does not affect the determination of l-tryptophan. Various influences were studied and a possible mechanism of perturbation to the B-Z oscillator by tryptophan was also discussed. Spectrophotometry and fluorescence spectrophotofluorimetry were used for comparision and confirmation of the results.     

A novel lipopolysaccharide-antagonizing aptamer protects mice against endotoxemia

A growing number of researchers have recognized the importance of using lipopolysaccharide (LPS) as target for the prevention and treatment of sepsis. However, no drugs targeting LPS have been applied clinically. In this study, LPS-inhibiting aptamers were screened by Systematic Evolution of Ligands by Exponential Enrichment (SELEX), and their therapeutic effects for experimental sepsis were observed. After 12 rounds of screening, 46 sequences were obtained. Primary structure analysis indicated that they had identical sequences, partly conserved sequences, or non-conserved sequences. Secondary structure analysis showed these sequences usually contained hairpin or stem-loop structures. Aptamer 19 significantly decreased NF-κB activation of monocytes challenged by LPS and reduced the IL-1 and TNF-α concentration in the media of LPS-challenged monocytes. Furthermore, aptamer 19 significantly increased the survival rate of mice with endotoxemia. The results suggest that a novel LPS antagonizing aptamer was obtained by SELEX, which successfully treated experimental sepsis.