Role of the indoleamine-2,3-dioxygenase/kynurenine pathway of tryptophan metabolism in behavioral alterations in a hepatic encephalopathy rat model.

BACKGROUND: This study aims to explore the role of indoleamine-2,3-dioxygenase (IDO)/kynurenine (KYN) pathway of tryptophan (TRY) metabolism in behavioral alterations observed in hepatic encephalopathy (HE) rats. METHODS: Expression levels of proinflammatory cytokines were tested by QT-PCR and ELISA, levels of IDOs were tested by QT-PCR and Western blot, and levels of 5-hydroxytryptamine (5-HT), KYN, TRY, 3-hydroxykynurenine (3-HK), and kynurenic acid (KA) in different brain regions were estimated using HPLC. Effects of the IDO direct inhibitor 1-methyl-L-tryptophan (1-MT) on cognitive, anxiety, and depressive-like behavior were evaluated in bile duct ligation (BDL) rats. RESULTS: Increased serum TNF-α, IL-1ß, and IL-6 levels were shown in rats 7 days after BDL, and these increases were observed earlier than those in the brain, indicating peripheral immune activation may result in central upregulation of proinflammatory cytokines. Moreover, BDL rats showed a progressive decline in memory formation, as well as anxiety and depressive-like behavior. Further study revealed that IDO expression increased after BDL, accompanied by a decrease of 5-HT and an increase of KYN, as well as abnormal expression of 3-HK and KA. The above results affected by BDL surgery were reversed by IDO inhibitor 1-MT treatment. CONCLUSION: Taken together, these findings indicate that (1) behavioral impairment in BDL rats is correlated with proinflammatory cytokines; (2) TRY pathway of KYN metabolism, activated by inflammation, may play an important role in HE development; and (3) 1-MT may serve as a therapeutic agent for HE.

Anti-ICAM-1 antibody-modified nanostructured lipid carriers: a pulmonary vascular endothelium-targeted device for acute lung injury therapy.

BACKGROUND: Acute lung injury (ALI) is a life-threatening clinical syndrome without effective treatment. Targeting delivery of glucocorticoid to lung shows potential efficacy for ALI based on their anti-inflammatory and anti-fibrotic properties, breaking through their clinical application limitation due to systemic side effects. This work was aimed to establish lung-targeted dexamethasone (DEX) loaded nanostructured lipid carriers (NLCs) with opposite surface charge and investigate their therapeutic effects on lipopolysaccharide (LPS)-induced ALI mice. RESULTS: The diameter of anionic anti-intercellular adhesion molecule 1 (anti-ICAM-1) antibody-conjugated DEX-loaded NLCs (ICAM/DEX/NLCs) and the cationic ones with octadecylamine (ODA) modification (ICAM/DEX/ODA-NLCs) was about 249.9 and 235.9 nm. The zeta potential of ICAM/DEX/NLCs and ICAM/DEX/ODA-NLCs was about - 30.3 and 37.4 mV, respectively. Relative to the non-targeted control and ICAM/DEX/ODA-NLCs, ICAM/DEX/NLCs exhibited higher in vitro cellular uptake in LPS-activated human vascular endothelial cell line EAhy926 after CAM-mediated endocytosis, and stronger in vivo pulmonary distribution in the ALI model mice. In vivo i.v. administration of ICAM/DEX/NLCs significantly attenuated pulmonary inflammatory cells infiltration, and the production of pro-inflammatory cytokine TNF-α and IL-6 in ALI mice. H&E stain also revealed positive histological improvements by ICAM/DEX/NLCs. CONCLUSIONS: ICAM/DEX/NLCs may represent a potential pulmonary endothelium targeted device, which facilitate translation of DEX into clinical ALI treatment.

The effects of fisetin on lipopolysaccharide-induced depressive-like behavior in mice.

Major depressive disorder (MDD) involves a series of pathological changes including the inflammation and increased cytokine levels. Fisetin, a natural flavonoid, has anti-inflammatory and antioxidant, and also has been shown in our previous studies to exert anti-depressant-like properties. The present study aimed to investigate the effect of fisetin on lipopolysaccharide (LPS)-induced depressive-like behavior and inflammation in mice. The results suggested that the immobility time in the forced swimming test (FST) and tail suspension test (TST) were increased at 6 h, 12 h and 24 h after LPS injection (0.83 mg/kg). However, only the group of 24 h treatment did not show any effect on locomotion counts. Pretreatment with fisetin at doses of 20, 40 and 80 mg/kg (p.o.) for 7 days reversed LPS-induced alterations of the immobility time in both of these two tests. Further neurochemical assays suggested that pretreatment with fisetin reversed LPS-induced overexpression of pro-inflammatory cytokine (IL-1ß, IL-6 and TNF-α) in the hippocampus and the prefrontal cortex (PFC). Moreover, higher dose of fisetin effectively antagonized iNOS mRNA expression and nitrite levels via the modulation of NF-κB in the hippocampus and PFC. Taken together, fisetin may be an effective therapeutic agent for LPS-induced depressive-like behaviors, which is due to its anti-inflammatory property.

Structure-based design of novel chemical modification of the 3'-overhang for optimization of short interfering RNA performance.

Short interfering RNAs (siRNAs) are broadly used to manipulate gene expression in mammalian cells. Although chemical modification is useful for increasing the potency of siRNAs in vivo, rational optimization of siRNA performance through chemical modification is still a challenge. In this work, we designed and synthesized a set of siRNAs containing modified two-nucleotide 3'-overhangs with the aim of strengthening the interaction between the 3'-end of the siRNA strand and the PAZ domain of Ago2. Their efficiency of binding to the PAZ domain was calculated using a computer modeling program, followed by measurement of RNA-Ago2 interaction in a surface plasmon resonance biochemical assay. The results suggest that increasing the level of binding of the 3'-end of the guiding strand with the PAZ domain, and/or reducing the level of binding of the sense strand through modifying the two-nucleotide 3'-overhangs, affects preferential strand selection and improves siRNA activity, while we cannot exclude the possibility that the modifications at the 3'-end of the sense strand may also affect the recognition of the 5'-end of the guiding strand by the MID domain. Taken together, our work presents a strategy for optimizing siRNA performance through asymmetric chemical modification of 3'-overhangs and also helps to develop the computer modeling method for rational siRNA design.

An efficient synthesis of novel dispirooxindole derivatives via one-pot three-component 1,3-dipolar cycloaddition reactions.

A series of novel dispirooxindoles have been synthesized through three-component 1,3-dipolar cycloaddition of azomethine ylides generated in situ by the decarboxylative condensation of isatin and an α-amino acid with the dipolarophile 5-benzylidene-1,3-dimethylpyrimidine-2,4,6-trione. This method has the advantages of mild reaction conditions, high atom economy, excellent yields, and high regio- and stereo-selectivity.

(Z)-Methyl 3-(2,4-dichloro-phen-yl)-3-hy-droxy-acrylate.

The mol-ecular structure of the title compound, C(10)H(8)Cl(2)O(3), exists in a cis-enol form, which is stabilized by a strong intra-molecular O-H⋯O hydrogen bond. In the crystal, C-H⋯O inter-actions generate zigzag chains along the c axis which are, in turn, linked by further C-H⋯O inter-actions into sheets parallel to (100).

Design, synthesis, and evaluation of 8-aminoquinoline-melatonin derivatives as effective multifunctional agents for Alzheimer's disease.

Background: Alzheimer's disease (AD) is thought to be a complex, multifactorial syndrome with many related molecular lesions contributing to its pathogenesis. Thus, multi-target-directed ligands are considered an effective way of treating AD. This study sought to evaluate 8-aminoquinoline-melatonin derivatives as effective multifunctional agents for AD. Methods: Thioflavin-T fluorescence assays were used to detect the inhibitory potency of 8-aminoquinoline-melatonin hybrids (a1-a5, b1-b5, and c1-c5) on self- and acetylcholinesterase (AChE)-induced amyloid-ß (Aß) aggregation. The AChE and butyrylcholinesterase (BuChE) inhibitory potency within the compounds was evaluated by Ellman's assays. Methyl thiazolyl tetrazolium (MTT) assays were performed to evaluate the cytotoxicity of the compounds to C17.2 cells. MTT assay was used to detect the cell viability of HT22 cells to evaluate the antioxidant effect of the compounds. Metal chelation property was measured by ultraviolet-visible spectrophotometry. Results: Compounds c3 and c5 had superior inhibitory activity against self-induced Aß aggregation (with inhibitory rates of 41.4±2.1 and 25.5±3.2 at 10 µM, respectively) compared to the other compounds. Compounds in the carbamate group (i.e., a4, a5, b4, b5, c4, and c5) showed significant BuChE inhibitory activity and excellent selectivity over AChE. Most of the compounds exhibited low cytotoxicity in the C17.2 cells. Notably, a2, a3, b2, and b3 and series c (c1-c5) exhibited strong protective effects. Additionally, a3 and c1 specifically chelated with copper ions. Conclusions: Taking all of the promising results together, 8-aminoquinoline-melatonin hybrids can serve as lead molecules in the further development of new multi-functional anti-AD agents.

Synthesis and in-vitro antibacterial activity of 7-(3-aminopyrrolo[3,4-c]pyrazol-5(2H,4H,6H)-yl)-6-fluoro-4-oxo-1,4-dihydroquinoline-3-carboxylic acid derivatives.

A series of novel 7-(3-aminopyrrolo[3,4-c]pyrazol-5(2H,4H,6H)-yl)-6-fluoro-4-oxo-1,4-dihydroquinoline-3-carboxylic acid derivatives was designed, synthesized and characterized by (1)H-NMR, MS and HRMS. These fluoroquinolones were evaluated for their in-vitro antibacterial activity against representative Gram-positive and Gram-negative strains. Generally, all of the target compounds display rather weak potency against the tested Gram-negative strains, but most of them exhibit good potency in inhibiting the growth of S. aureus including methicillin-resistant Staphylococcus aureus (MRSA) and Staphylococcus epidermidis including methicillin-resistant S. epidermidis (MRSE) (MIC: 0.125-8 µg/mL). In particular, the compound 9g is 2 to 32 fold more potent than gemifloxacin (GM), moxifloxacin (MX), gatifloxacin (GT), and levofloxacin (LV) against S. pneumoniae 08-3, K. pneumoniae 09-23, and P. aeruginosa ATCC27853, 4 to 32 fold more potent than MX, GM, and LV against K. pneumoniae 09-21, and more active than or comparable to the four reference drugs against P. aeruginosa 09-32.

Lipopolysaccharide-induced depression is associated with estrogen receptor-α/SIRT1/NF-κB signaling pathway in old female mice.

The present study aims to investigate the influence of sex/age on depressive-like behaviors in lipopolysaccharide (LPS)-challenged mice model, and explore the underlying mechanisms. Tail suspension test and forced swimming test were used to evaluate the depressive-like behaviors. SIRT1 mRNA expression was assessed by PCR. Levels of 17ß-estradiol (E2), SIRT1, NF-κB, tumor necrosis factor α (TNF-α), interleukin 1ß (IL-1ß) and interleukin 6 (IL-6) were detected by enzyme linked immunosorbent assay (ELISA). In the behavior tests, under the same LPS stimulation, significant depressive-like behavior was observed in young male mice but not in young female mice, however, female mice were more likely to be depressed than male mice in the old age. Moreover, we found age-related depression difference existed only in female mice. In the experiments of mechanism exploration in old female mice, E2 improved LPS-induced depressive-like behavior, and simultaneously elevated SIRT1 levels and downregulated expressions of NF-κB and inflammatory cytokines in the hippocampus and frontal cortex. Interestingly, ERα inhibition, not ERß inhibition, abolished E2's function. Additionally, SIRT1 antagonist also reversed E2's effects on depressive-like behavior and the expressions of NF-κB and inflammatory cytokines. These results suggested that E2 could protect the old female mice from depression via E2/ERα/SIRT1/NF-κB signaling pathway. In other words, LPS-induced depression was associated with ER-α/SIRT1/NF-κB signaling pathway in old female mice. By comparing the results of mechanism exploration in old male mice and old female mice and the different expression levels of E2, SIRT1, NF-κB and inflammatory cytokines in young female mice and old female mice, we speculate that the age or gender-related depression difference may be associated with the different activation levels of the ERα/SIRT1/NF-κB signaling pathway.

Ferulic acid improves motor function induced by spinal cord injury in rats via inhibiting neuroinflammation and apoptosis.

PURPOSE: To investigate the effect of ferulic acid (FA) on spinal cord injury (SCI)-induced motor dysfunction and to explore the possible pharmacological mechanisms. METHODS: Adult male Wistar rats were used in our study. SCI was achieved by clipping the spinal cord T9 of the rat by a vascular clip for 2 minutes. The motor function of the rat was evaluated by Basso, Beattie, and Bresnahan scoring method (BBB) and inclined plane test. Hematoxylin and eosin (HE) staining, NISSL staining, and transmission electron microscopic examination were used to evaluate alterations at the histological level. Polymerase chain reaction (PCR), Western blots, and enzyme-linked immunosorbent assays (ELISA) were employed in biochemical analysis. RESULTS: The BBB score and inclined plane test score significantly decreased after SCI surgery, whereas chronic FA treatment (dose of 90 mg/kg, i.g.) for 28 days improved SCI-induced motor dysfunction. HE staining showed that SCI surgery induced internal spinal cord edema, but the structural changes of the spinal cord could be reversed by FA treatment. NISSL staining and transmission electron microscopic examination confirmed the improvement of the effect of FA on the injury site. In the biochemical analysis, it could be found that FA inhibitedSCI-induced mRNA and protein overexpression of pro-inflammatory cytokines (IL-1ß, IL-6, TNF-α), as well as iNOS and COX-2 via the modulation of NF-κB level in the spinal cord of SCI rat. Moreover, the SCI-induced decrease of Bcl-2/Bax ratio was also reversed by FA treatment. However, the effect of FA on the expression of Beclin-1 was not statistically significant. CONCLUSIONS: FA showed a therapeutic effect on SCI, which may be associated with the regulation of neuroinflammation and apoptosis.

Irisin protects female mice with LPS-induced endometritis through the AMPK/NF-κB pathway.

OBJECTIVES: This research was designed to determine the role of irisin in lipopolysaccharide (LPS)-induced endometritis in female mice. MATERIALS AND METHODS: Animals were randomly assigned into sham, sham + irisin, LPS, LPS + irisin (0.1, 1, 10 µg/kg), and LPS + irisin + compound C groups. Histological features and expression of AMPK, NF-κB, inflammatory mediators, and oxidative stress markers were compared among different groups. RESULTS: The results showed that LPS resulted in obvious uterus damage, meanwhile, the inflammatory mediators (COX-2, iNOS, IL-1ß, IL-6, and TNF-α), as well as NF-κB in the uterine tissue, were significantly increased and the level of adenosine monophosphate-activated protein kinase (AMPK) was reduced. Nevertheless, pretreatment with irisin reversed the phenomena caused by LPS. Interestingly, compound C (AMPK inhibitor) abolished irisin's effects on the uterus, which suggested that irisin's beneficial function was achieved through regulating the AMPK-NF-κB pathway. Moreover, LPS-induced alterations of oxidative factors (MnSOD, GSH, and MDA) were reversed significantly by pretreatment with irisin. This data indicated irisin's beneficial function was also related to antioxidation besides anti-inflammation. CONCLUSION: Our study implies that irisin is a potential therapeutic agent for endometritis.

Indoleamine-2,3-Dioxygenase Mediates Emotional Deficits by the Kynurenine/Tryptophan Pathway in the Ethanol Addiction/Withdrawal Mouse Model.

OBJECTIVE: Our study was designed to investigate whether the indoleamine-2,3-dioxygenase (IDO)-mediated kynurenine/tryptophan (KYN/TRP) pathway participates in the development of emotional deficits from ethanol addiction/withdrawal mice. METHODS: The expression of proinflammatory factors, including tumor necrosis factor α (TNF-α), interleukin-1ß (IL-1ß), and interleukin-6 (IL-6), was tested by enzyme-linked immunosorbent assay (ELISA). The IDO levels in the hippocampus, cerebral cortex, and amygdala were measured by polymerase chain reaction (PCR) and western blot, and the neurotransmitters were tested by high performance liquid chromatography (HPLC). Emotional deficits of mice were evaluated by behavioral tests. RESULTS: Expression levels of inflammatory factors (TNF-α, IL-1ß, and IL-6) were increased in mice after 4 weeks of alcohol exposure. As for indoleamine 2,3-dioxygenase (IDO) expression, only the subtype IDO1 was found to increase at both mRNA level and protein level in all the tested brain regions of ethanol addiction/withdrawal mice. In behavioral tests, mice exposed to alcohol showed gradually declined memory function accompanied by anxiety-like and depressive-like behaviors. Meanwhile, increased expression of KYN, decreased expression of 5-HT, and abnormal expression of 3-HK and KA were found in the hippocampus, cerebral cortex, and amygdala of ethanol addiction/withdrawal mice. Interestingly, the IDO1 inhibitor, 1-methyl-L-tryptophan (1-MT), reversed all above alterations induced by ethanol in mice. CONCLUSION: Our results suggested that the TRP/KYN pathway, medicated by IDO1, in the hippocampus, cerebral cortex, and amygdala, plays an important role in the development of emotional deficits caused by ethanol addiction and withdrawal.

Irisin Protects Against Motor Dysfunction of Rats with Spinal Cord Injury via Adenosine 5'-Monophosphate (AMP)-Activated Protein Kinase-Nuclear Factor Kappa-B Pathway.

The aim of the present research was to investigate the effects of irisin, a skeletal muscle-derived myokine, on spinal cord injury (SCI) in rats and explore the possible mechanisms. SCI model was constructed in male SD rats. The effects of irisin on SCI rats were assessed via behavior tests including Basso, Beattie, and Bresnahan (BBB) scoring method and inclined plane test, followed by histomorphology tests including HE staining, Nissl staining, and transmission electron microscope examination. Biochemical analyses including PCR, Western blots and ELISA were employed to further evaluate the changes at molecular level of SCI rats. In addition, lipopolysaccharide (LPS)-induced cell damage model was established in PC12 cells to verify the mechanism of irisin's effect on nerve cells in vitro. Results showed that the BBB score and the angle of incline significantly decreased after SCI surgery, however, chronic irisin treatment improved SCI-induced motor dysfunction. HE and Nissl staining assays showed that SCI surgery induced histological injury of spinal cord, which could be reversed by irisin treatment. Morphological abnormality of nerve cells caused by SCI also could be alleviated by irisin. Further biochemical analyses showed that irisin inhibited SCI-induced overexpression of Interleukin-1ß (IL-1ß), Interleukin- 6 (IL-6), tumor necrosis factor alpha (TNF-α), inducible nitricoxidesynthase (iNOS) and Cyclooxygenase-2 (COX-2)], as well as nuclear factor kappa-B (NF-κB)p65 in rats, and the positive function of irisin could be reversed by Compound C treatment. In our in vitro study, LPS-induced declines of cell viability and neurite length of PC12 cell were inhibited by irisin treatment, and irisin inhibited LPS-induced overexpression of NF-κBp65, IL-1ß, IL-6, TNF-α, iNOS and COX-2. These changes could be reversed by activated protein kinase (AMPK) siRNA pre-treatment. Taken together, irisin could protect the rats from SCI, and its protection is associated with the regulation of adenosine 5'-monophosphate-activated protein kinase (AMPK)- NF-κB signaling pathway.

Trans-astaxanthin attenuates lipopolysaccharide-induced neuroinflammation and depressive-like behavior in mice.

Mounting evidence supports that inflammation and increased cytokine levels are associated with depression-like symptoms and neuropsychological disturbances in humans. Trans-astaxanthin has anti-inflammatory and antioxidative activity, also has the ability to cross the blood-brain barrier in rodents. Here, we investigated the effects of trans-astaxanthin on lipopolysaccharide (LPS)-induced depressive-like behavior in mice. In both the forced swimming test (FST) and tail suspension test (TST), the immobility time was increased when mice were administrated with a single dose of LPS (0.83mg/kg, i.p.). However, this alteration can be reversed by pretreatment of trans-astaxanthin at doses of 20, 40 and 80mg/kg (p.o.) for 7 days. Further neurochemical assays suggested that LPS-induced overexpression of pro-inflammatory cytokines (IL-1ß, IL-6 and TNF-α) in the hippocampus and the prefrontal cortex (PFC) can also be reversed by trans-astaxanthin treatment. Moreover, trans-astaxanthin at 80mg/kg was demonstrated to effectively antagonize iNOS, nNOS and COX-2 expression, both at mRNA and protein levels, nitric oxide (NO) levels, via regulating NF-κB in the hippocampus and PFC. Taken together, trans-astaxanthin may serve as an effective therapeutic agent for LPS-induced depressive-like behavior via its potent anti-inflammatory property.

Ferulic acid improves motor function induced by spinal cord injury in rats via inhibiting neuroinflammation and apoptosis

ABSTRACT Purpose To investigate the effect of ferulic acid (FA) on spinal cord injury (SCI)-induced motor dysfunction and to explore the possible pharmacological mechanisms. Methods Adult male Wistar rats were used in our study. SCI was achieved by clipping the spinal cord T9 of the rat by a vascular clip for 2 minutes. The motor function of the rat was evaluated by Basso, Beattie, and Bresnahan scoring method (BBB) and inclined plane test. Hematoxylin and eosin (HE) staining, NISSL staining, and transmission electron microscopic examination were used to evaluate alterations at the histological level. Polymerase chain reaction (PCR), Western blots, and enzyme-linked immunosorbent assays (ELISA) were employed in biochemical analysis. Results The BBB score and inclined plane test score significantly decreased after SCI surgery, whereas chronic FA treatment (dose of 90 mg/kg, i.g.) for 28 days improved SCI-induced motor dysfunction. HE staining showed that SCI surgery induced internal spinal cord edema, but the structural changes of the spinal cord could be reversed by FA treatment. NISSL staining and transmission electron microscopic examination confirmed the improvement of the effect of FA on the injury site. In the biochemical analysis, it could be found that FA inhibitedSCI-induced mRNA and protein overexpression of pro-inflammatory cytokines (IL-1β, IL-6, TNF-α), as well as iNOS and COX-2 via the modulation of NF-κB level in the spinal cord of SCI rat. Moreover, the SCI-induced decrease of Bcl-2/Bax ratio was also reversed by FA treatment. However, the effect of FA on the expression of Beclin-1 was not statistically significant. Conclusions FA showed a therapeutic effect on SCI, which may be associated with the regulation of neuroinflammation and apoptosis.