Untargeted metabolomics analysis of rat hippocampus subjected to sleep fragmentation

Sleep fragmentation (SF) commonly occurs in several pathologic conditions and is especially associated with impairments of hippocampus-dependent neurocognitive functions. Although the effects of SF on hippocampus in terms of protein or gene levels were examined in several studies, the impact of SF at the metabolite level has not been investigated. Thus, in this study, the differentially expressed large-scale metabolite profiles of hippocampus in a rat model of SF were investigated using untargeted metabolomics approaches. Forty-eight rats were divided into the following 4 groups: 4-day SF group, 4-day exercise control (EC) group, 15-day SF group, and 15-day EC group (n = 12, each). SF was accomplished by forced exercise using a walking wheel system with 30-s on/90-s off cycles, and EC condition was set at 10-min on/30-min off. The metabolite profiles of rat hippocampi in the SF and EC groups were analyzed using liquid chromatography/mass spectrometry. Multivariate analysis revealed distinctive metabolic profiles and marker signals between the SF and corresponding EC groups. Metabolic changes were significant only in the 15-day SF group. In the 15-day SF group, L-tryptophan, myristoylcarnitine, and palmitoylcarnitine were significantly increased, while adenosine monophosphate, hypoxanthine, L-glutamate, L-aspartate, L-methionine, and glycerophosphocholine were decreased compared to the EC group. The alanine, aspartate, and glutamate metabolism pathway was observed as the common key pathway in the 15-day SF groups. The results from this untargeted metabolomics study provide a perspective on metabolic impact of SF on the hippocampus.Peer reviewe

Amyloid-β peptide induces oligodendrocyte death by activating the neutral sphingomyelinase–ceramide pathway

Amyloid-β peptide (Aβ) accumulation in senile plaques, a pathological hallmark of Alzheimer's disease (AD), has been implicated in neuronal degeneration. We have recently demonstrated that Aβ induced oligodendrocyte (OLG) apoptosis, suggesting a role in white matter pathology in AD. Here, we explore the molecular mechanisms involved in Aβ-induced OLG death, examining the potential role of ceramide, a known apoptogenic mediator. Both Aβ and ceramide induced OLG death. In addition, Aβ activated neutral sphingomyelinase (nSMase), but not acidic sphingomyelinase, resulting in increased ceramide generation. Blocking ceramide degradation with N-oleoyl-ethanolamine exacerbated Aβ cytotoxicity; and addition of bacterial sphingomyelinase (mimicking cellular nSMase activity) induced OLG death. Furthermore, nSMase inhibition by 3-O-methyl-sphingomyelin or by gene knockdown using antisense oligonucleotides attenuated Aβ-induced OLG death. Glutathione (GSH) precursors inhibited Aβ activation of nSMase and prevented OLG death, whereas GSH depletors increased nSMase activity and Aβ-induced death. These results suggest that Aβ induces OLG death by activating the nSMase–ceramide cascade via an oxidative mechanism

Scale-up study for ex-vivo expansion of allogeneic natural killer cells in stirred-tank bioreactor

Natural killer (NK) cells are a type of lymphocyte in the blood that are responsible for innate and adaptive immune response, and they mature in the liver and bone marrow. Being a key role in host defense system with direct and indirect killing of virus-infected cells or cancer cells, NK cell has been considered an attractive candidate for cancer therapy. Peripheral blood shows the low frequency of NK cells, so ex vivo expansion method is important to obtain sufficient NK cells for therapeutic use. Currently, we successfully developed bioreactor process for NK cell expansion on lab-scale. Stirred-tank bioreactor could be considered as optimal alternative system for large-scale NK cell expansion compared with other ones because it is automated, less labor intensive, scalable, well-controlled and cost-effective. In bioreactor process, agitation is one of important parameters for NK cell expansion because it is necessary to provide homogenous culture conditions. So we defined effects of agitation in bioreactor and figured out an optimum condition. After that scale-up studies were carried out with manufacturing-scale bioreactor based on these results. The results in terms of growth rate, viability cytotoxicity and purity, were comparable with lab-scale

In vitro antioxidant and anti-adipogenic effects of slendesta, standard potato extracts containing 5% protease inhibitor II

Background: The objective of the present study is to observe the anti-adipogenic effects of Slendesta (SLD), a standard potato protein extracts containing 5% potato protease inhibitor II (PI2) on the 3T3-L1 preadipocytes which are able to differentiate into mature adipocytes and accumulate lipids, as an obesity model with cytotoxicity and antioxidant effects.Materials and Methods: The cytotoxicity of SLD was observed against 3T3-L1 preadipocyte cell line by MTT assay, and also antiadipogenic effects were observed through lipid accumulation assay during 3T3-L1 differentiation as comparing with N-Acetyl-Lcysteine (NAC). In addition, antioxidant effects of SLD were detected by free radical scavenging capacity and superoxide dismutase (SOD)-like activity as comparing with ascorbic acid.Results: The SLD showed obvious cytotoxicity against 3T3-L1 pre-adipocyte cell line at higher concentrations, from 1.5 mg/ml for 72 h treatment, and the cytotoxic IC50 of SLD after 24, 48 and 72 h treatment times were detected as 10.11 ± 0.67, 5.71 ± 0.37 and 5.34 ± 0.21 mg/ml, respectively. The SLD also concentration-dependently inhibited the lipid accumulations formatted during 3T3-L1 cell differentiations. The adipogenic specific genes including PPARγ, C/EBPα, C/EBPβ and leptin were found to be reduced in SLD and NAC-treated cells compared to control cells. Furthermore, the SLD effectively showed DPPH radical scavenging activity (IC50 = 161.98 ± 64.65 μg/ml) and SOD-like effects (IC50 = 284.54 ± 54.47 μg/ml), and the cellular ROS was significantly inhibited in the SLD-treated cells compared to control cells.Conclusion: The results suggest that SLD effectively inhibit the differentiations of 3T3-L1 preadipose cell probably through antioxidant activities and direct cytotoxicity in case of higher concentration, along with satiety effects mediated by increases of circulating cholecystokinin. These findings are considered as direct evidences that SLD may serve as a predictable functional ingredient for obesity as an alternative therapy.Key words: Slendesta, potato protease inhibitor II, 3T3-L1 cell, cytotoxicity, anti-adipogenic effects, antioxidant effects

An Unusual Case with Membranous Lipodystrophy in a Hypertensive Patient with Transepidermal Elimination

Membranous lipodystrophy represents a peculiar type of fat necrosis that is present in patients with various types of skin disease. It is characterized by the presence of microcysts and macrocysts and is lined by amorphous eosinophilic material with a crenelated arabesque appearance. These findings have been associated with lupus erythematosus, diabetes mellitus, erythema nodosum, trauma, etc. We report a case of a 43-year-old woman who had a red to purple asymptomatic indurated plaque, approximately seven cm in diameter and on the left arm. She was a chronic hepatitis B antigen carrier and had hypertension for four years. Histopathology of the biopsied lesion showed transepidermal elimination of altered collagen and elastic fibers, as well as membranous lipodystrophy changes. There were hypertensive vascular changes including lymphohistiocytic infiltration around the vascular wall, swelling of endothelial cells, increased thickness of the vascular walls, and narrowing of the lumen. We report a case showing transepidermal elimination with membranous lipodystrophy. We carefully suggest that the secondary phenomenon of transepidermal elimination was associated with membranous lipodystrophy and degenerate connective tissues

Differential Proteome Profiling Using iTRAQ in Microalbuminuric and Normoalbuminuric Type 2 Diabetic Patients

Diabetic nephropathy (DN) is a long-term complication of diabetes mellitus that leads to end-stage renal disease. Microalbuminuria is used for the early detection of diabetic renal damage, but such levels do not reflect the state of incipient DN precisely in type 2 diabetic patients because microalbuminuria develops in other diseases, necessitating more accurate biomarkers that detect incipient DN. Isobaric tags for relative and absolute quantification (iTRAQ) were used to identify urinary proteins that were differentially excreted in normoalbuminuric and microalbuminuric patients with type 2 diabetes where 710 and 196 proteins were identified and quantified, respectively. Some candidates were confirmed by 2-DE analysis, or validated by Western blot and multiple reaction monitoring (MRM). Specifically, some differentially expressed proteins were verified by MRM in urine from normoalbuminuric and microalbuminuric patients with type 2 diabetes, wherein alpha-1-antitrypsin, alpha-1-acid glycoprotein 1, and prostate stem cell antigen had excellent AUC values (0.849, 0.873, and 0.825, resp.). Moreover, we performed a multiplex assay using these biomarker candidates, resulting in a merged AUC value of 0.921. Although the differentially expressed proteins in this iTRAQ study require further validation in larger and categorized sample groups, they constitute baseline data on preliminary biomarker candidates that can be used to discover novel biomarkers for incipient DN