Structure and energetics of serum protein complex of tea adulterant dye Bismarck brown Y using experimental and computational methods.

Tea, a widely consumed aromatic beverage, is often adulterated with dyes such as Bismarck brown Y (C.I. 21000) (BBY), Prussian blue, and Plumbago, which pose potential health risks. The objective of this study is to analyze how the food dye BBY interacts with serum protein, bovine serum albumin (BSA). This study investigated the BBY-BSA interaction at the molecular level. Fluorescence spectroscopy results showed that the quenching of BSA by BBY is carried out by dynamic quenching mechanism. The displacement assay and molecular docking studies revealed that BBY binds at the flavanone binding site of BSA with hydrophobic interactions. Circular Dichroism results indicate the structural stability of the protein upon BBY binding. Molecular dynamics simulations demonstrated the stability of the complex in a dynamic solvent system, and quantum mechanics calculations showed slight conformational changes of the diaminophenyl ring due to increased hydrophobic interaction. The energetics of gas phase optimized and stable MD structures of BBY indicated similar values which further confirmed that the conformational changes were minor, and it also exhibited a moderate binding with BSA as shown by the MM/PBSA results. This study enhances our understanding of the molecular-level interactions between BBY and BSA, emphasizing the critical role of hydrophobic interactions.

Immune-biochemical response and immune gene expression profiling of Labeo rohita fingerlings fed with ethanolic tea leaf extracts and its survivability against Aeromonas hydrophila infection.

The present study was conducted to evaluate the immunostimulatory effect of tea leaf extract (Camellia sinensis) on Labeo rohita and its resistance against Aeromonas hydrophila infection. The ethanolic extract of green tea (GTEE) was found to be the most potent as compared to other solvent extract which was used for further study. It was used to evaluate immune-biochemical response of L. rohita fingerlings, fed with tea leaf extract (control- 0.0%, 0.2% (T1), 0.4% (T2), 0.8% (T3) and 1% (T4) of GTEE kg-1 feed). Different biochemical parameters like glucose, ALP, GPT, GOT, and immunological parameters like lysozyme activity, NBT, anti-protease activity, myeloperoxidase activity, plasma protein, and immune relevant genes (IL-10, C3, Lysozyme G type and iNOS) expressions were carried out. The immunological parameters such as lysozyme activity, NBT and myeloperoxidase activity showed significantly high value once fed with GTEE incorporated diets. Significant up-regulation of immune genes indicated the enhancement of immune response at molecular level. The biochemical parameters were found to be significantly decreasing, indicating that the extract had hepato-protective effect and can help to overcome stress. The fish, fed with GTEE incorporated diets, showed resistance against A. hydrophila when compared with the control group. 0.2% GTEE showed the highest post-challenged survival (76.67%). From the present study, it is concluded that GTEE @ 0.2% can be used as potent immunostimulant as a sustainable alternative prophylactic and therapeutic agent in aquaculture.

The Influence of Rumen-Protected Choline and α-tocopherol Supplementation on Early Lactating Dairy Cows Metabolism.

It is well documented that choline is known as one of the essential ingredients of phospholipids. Choline acts as a determinative element for appropriate cell membrane functions. On the other hand α-tocopherol (Vit E) is a fat-soluble vitamin. This vitamin acts as a strong antioxidant in the living body's defense system against oxidative stress. Lipid peroxidation in peripartum and early lactating cows is significantly increased while the level of serum Vit E is decreases dramatically. These concomitant physiological changes demonstrate a higher level of oxidative stress subsequently leads to serious health issues in dairy cows. Therefore, the present research was designed to investigate the following items in dairy cattle: 1) evaluation of the possible changes in serum protein fractions, and 2) comparing the oxidative status of orally RPC and vitamin E supplementation in dairy cows in early lactation period. In the current study 30 early lactating primiparous and multiparous Holstein cows (body condition score (BCS)=2.51 ± 0.10) were used beginning five weeks postpartum. All the animals were randomly divided in to three groups (n=10) (number of lactation=2.61). The animals were randomly assigned to receive one of the following treatments. Group 1 served as control group were not received any supplement. The second group was supplemented with 90 g/d of RPC (Reashre Choline, Balchem, USA). The third group was administrated 4400 IU/d vitamin E (Roche, Vitamins Ltd; Switzerland). In the current study, serum protein electrophoresis showed four main fractions as follows: albumin, α-globulin, ß-globulin, and γ-globulin. The recorded data showed that the percentages of albumin and γ-globulin fractions were higher in treated groups compared to the control group. In the animals supplementing with RPC and vitamin E the percentages of serum albumin increased to the value of 37. 70±1.63 and 38.21±1.28 respectively compare to the control group (34.69±1.21), which were significant (P<0.05).

Pharmacokinetics, plasma protein binding, and metabolism of a potential natural chemosensitizer from Marsdenia tenacissima in rats.

ETHNOPHARMACOLOGICAL RELEVANCE: Marsdenia tenacissima (Roxb.) Wight et Arn is a medicinal plant mainly distributed in southwest China. It is used in folk medicine for the treatment of tumors and is synergistic with chemotherapies. In our previous study, 11α-O-2-methybutyryl-12ß-O-tigloyl-tenacigenin B (MT2), a main steroid aglycone isolated from the total aglycones of M. tenacissima, significantly enhanced the in vivo antitumor effect of paclitaxel in mice bearing human tumor xenografts, showing its potential as a chemosensitizer. However, the pharmacokinetic characteristics, plasma protein binding rate, and metabolic profile of MT2 remain unclear. AIM OF THE STUDY: To elucidate the pharmacokinetic characteristics, plasma protein binding rate, and metabolic profile of MT2 in rats. MATERIALS AND METHODS: MT2 in rat plasma and phosphate-buffered saline was quantified using ultra performance liquid chromatography tandem mass spectrometry (UPLC-MS/MS) method, while the MT2 metabolites in rat liver microsomes were analyzed using UPLC-triple time-of-flight MS/MS. RESULTS: For intravenously administered MT2, the maximum plasma concentration and the area under the plasma concentration-time curve indicated dose dependency, while the elimination half-life time, the mean residence time, apparent volume of distribution and total apparent clearance values remained relatively unchanged in both the 5 mg/kg and 10 mg/kg groups. For orally administered MT2, the bioavailability was 1.08-1.11%. In rat plasma, MT2 exhibited a protein binding rate of 93.84-94.96%. In rat liver microsomes, MT2 was metabolized by oxidation alone or in combination with demethylation, and five MT2 metabolites were identified. CONCLUSION: MT2 has low oral bioavailability and a high plasma protein binding rate in rats. After administration, MT2 is transformed into oxidative metabolites in the liver. To achieve a high blood concentration of MT2, it should be administered intravenously. These findings would serve as a reference for further MT2-based pharmacological study and drug development.

Two weeks of moderate intensity locomotor training increased corticosterone concentrations but did not alter the number of adropin-immunoreactive cells in the hippocampus of diabetic type 2 and control rats.

Adropin (ADR) plays a role in metabolism regulation and its alterations in obesity and diabetes have been found. Treatment with ADR was beneficial in metabolic diseases, and physical exercise increased ADR concentrations in obese patients. However, data on the distribution of ADR in the brain are sparse. The role of metabolic status and physical exercise on its expression in the brain is undiscovered. We hypothesized that diabetes type 2 (DM2) and/or exercise will alter number of ADR-immunoractive (-ir) cells in the rat brain. Animals were divided into groups: diabetes type 2 (receiving high-fat diet and injections of streptozotocin) and control (fed laboratory chow diet; C). Rats were further divided into: running group (2 weeks of forced exercise on a treadmill) and non-running group. Body mass, metabolic and hormonal profiles were assessed. Immunohistochemistry was run to study ADR-ir cells in the brain. We found that: 1) in DM2 animals, running decreased insulin and increased glucose concentrations; 2) in C rats, running decreased insulin concentrations and had no effect on glucose concentration in blood; 3) running increased corticosterone (CORT) concentrations in DM2 and C rats; 4) ADR-ir cells were detected in the hippocampus and ADR-ir fibers in the arcuate nucleus of the hypothalamus, which is a novel location; 5) metabolic status and running, however, did not change number of these cells. We concluded that 2 weeks of forced moderate intensity locomotor training induced stress response present as increased concentration of CORT and did not influence number of ADR-ir cells in the brain.

Elucidation of plasma protein binding, blood partitioning, permeability, CYP phenotyping and CYP inhibition studies of Withanone using validated UPLC method: An active constituent of neuroprotective herb Ashwagandha.

ETHNOPHARMACOLOGICAL RELEVANCE: Withanone (WN), an active constituent of Withania somnifera commonly called Ashwagandha has remarkable pharmacological responses along with neurological activities. However, for a better understanding of the pharmacokinetic and pharmacodynamic behavior of WN, a comprehensive in-vitro ADME (absorption, distribution, metabolism, and excretion) studies are necessary. AIM OF THE STUDY: A precise, accurate, and sensitive reverse-phase ultra-performance liquid chromatographic method of WN was developed and validated in rat plasma for the first time. The developed method was successfully applied to the in-vitro ADME investigation of WN. MATERIAL AND METHODS: The passive permeability of WN was assayed using PAMPA plates and the plasma protein binding (PPB) was performed using the equilibrium dialysis method. Pooled liver microsomes of rat (RLM) and human (HLM) were used for the microsomal stability, CYP phenotyping, and inhibition studies. CYP phenotyping was evaluated using the specific inhibitors. CYP inhibition study was performed using specific probe substrates along with WN or specific inhibitors. RESULTS: WN was found to be stable in the simulated gastric and intestinal environment and has a high passive permeability at pH 4.0 and 7.0 in PAMPA assay. The PPB of WN at 5 and 20 µg/mL concentrations were found to be high i.e. 82.01 ± 1.44 and 88.02 ± 1.15%, respectively. The in vitro half-life of WN in RLM and HLM was found to be 59.63 ± 2.50 and 68.42 ± 2.19 min, respectively. CYP phenotyping results showed that WN was extensively metabolized by CYP 3A4 and1A2 enzymes in RLM and HLM. However, the results of CYP Inhibition studies showed that none of the CYP isoenzymes were potentially inhibited by WN in RLM and HLM. CONCLUSION: The in vitro results of pH-dependent stability, plasma stability, permeability, PPB, blood partitioning, microsomal stability, CYP phenotyping, and CYP inhibition studies demonstrated that WN could be a better phytochemical for neurological disorders.

Ultrafiltration isolation, structures and anti-tumor potentials of two arabinose- and galactose-rich pectins from leaves of Aralia elata.

Two novel arabinose- and galactose-rich pectic polysaccharides, AELP-B5 (Mw, 4.25 × 104 g/mol) and B6 (Mw, 1.56 × 104 g/mol), were rapidly obtained from the leaves of Aralia elata (Miq.) Seem. with anion resin and sequenced ultrafiltration membrane columns. The structural backbone and branched chains of AELP-B5 and B6 were preliminarily elucidated by mild acid hydrolysis with HILIC-ESI--MS/MS. The planar structures and spatial configurations were further identified using UPLC-QDa and GC-MS for compositions, Smith degradation and methylation analysis, FT-IR, NMR (1H/13C, DEPT, HSQC, HMBC, COSY, NOESY and TOCSY) and SEC-MALLS-RID. (1) AELP-B5 possessed →4GalA1→ as smooth regions (HG) and a repeating disaccharide moiety of →4GalA1→2Rha1→ as hairy regions (RG-I) with a 1:5 molar ratio, whereas AELP-B6 had a distinguishing 1:1 molar ratio between the HG and RG-I; (2) complex side chains were constituted of T-α-Araf, 1,3-α-Araf, 1,5-α-Araf, T-ß-Galp, 1,3-ß-Galp, 1,4-ß-Galp, 1,6-ß-Galp, 1,3,4-ß-Galp and 1,3,4,6-ß-Galp connected at C-4 of the rhamnosyl units in RG-I of AELP-B5 and B6; and (3) both possessed highly branched and compact coil conformations. The CCK-8 assay illustrated that AELP-B6 possessed higher cytotoxicity against HepG2 and HT-29 than that of AELP-B5. Surface plasmon resonance showed the binding affinity of AELP-B6 to galectin-3 (6.488 × 10-5 M) was about 10 times stronger than that of AELP-B5 (4.588 × 10-4 M). The above findings provide a molecular structure and bioactivity basis for future potential applications of AELP in the food and medical industries.

It's in Our Blood: A Glimpse of Personalized Medicine.

Recent advances in protein profiling technology has facilitated simultaneous measurement of thousands of proteins in large population studies, exposing the depth and complexity of the plasma and serum proteomes. This revealed that proteins in circulation were organized into regulatory modules under genetic control and closely associated with current and future common diseases. Unlike networks in solid tissues, serum protein networks comprise members synthesized across different tissues of the body. Genetic analysis reveals that this cross-tissue regulation of the serum proteome participates in systemic homeostasis and mirrors the global disease state of individuals. Here, we discuss how application of this information in routine clinical evaluations may transform the future practice of medicine.

Cell type-selective secretome profiling in vivo.

Secreted polypeptides are a fundamental axis of intercellular and endocrine communication. However, a global understanding of the composition and dynamics of cellular secretomes in intact mammalian organisms has been lacking. Here, we introduce a proximity biotinylation strategy that enables labeling, detection and enrichment of secreted polypeptides in a cell type-selective manner in mice. We generate a proteomic atlas of hepatocyte, myocyte, pericyte and myeloid cell secretomes by direct purification of biotinylated secreted proteins from blood plasma. Our secretome dataset validates known cell type-protein pairs, reveals secreted polypeptides that distinguish between cell types and identifies new cellular sources for classical plasma proteins. Lastly, we uncover a dynamic and previously undescribed nutrient-dependent reprogramming of the hepatocyte secretome characterized by the increased unconventional secretion of the cytosolic enzyme betaine-homocysteine S-methyltransferase (BHMT). This secretome profiling strategy enables dynamic and cell type-specific dissection of the plasma proteome and the secreted polypeptides that mediate intercellular signaling.

Blood Metabolic and Estradiol Level of Repeat Breeder and Fertile in Friesian Holstein Cross Breed Cows in the Tropic.

BACKGROUND AND OBJECTIVE: The repeat breeding becomes the major reproduction problem in tropic area especially in Indonesia. It relates to blood metabolic and hormonal level. This research was conducted to investigate the level of blood metabolic and estradiol between the repeat breeder Friesian Holstein Cross Breed Cows (FHCB) and the fertile FHCB cows. MATERIALS AND METHODS: Twenty FHCB cows in luteal phase on 2nd to 3rd lactation were used in this research. Group I consist of 10 repeat breeder FHCB cows and group II consist of 10 fertile FHCB cows. Blood samples were collected through jugular vein prior to feeding. The level of total protein, phosphorus, glucose, cholesterol and estradiol in serum were calculated. The data were analyzed by using the independent samples t-test for comparing the blood metabolic and estradiol between the repeat breeder FHCB cows and the fertile FHCB cows. RESULTS: The results showed that repeat breeder FHCB cows were found to have lower level of all of the parameters of blood metabolic and estradiol descriptively, however, only the serum cholesterol and serum phosphorus had the significant difference (p<0.05) between the repeat breeder FHCB cows and the fertile FHCB cows. CONCLUSION: It could be concluded that level of serum cholesterol and serum phosphorus played a role in repeat breeding.

In Vitro Hepatic Uptake in Human and Monkey Hepatocytes in the Presence and Absence of Serum Protein and Its In Vitro to In Vivo Extrapolation.

It is well documented that human hepatic clearance based on in vitro metabolism or transporter assays systematically resulted in underprediction; therefore, large empirical scalars are often needed in either static or physiologically based pharmacokinetic (PBPK) models to accurately predict human pharmacokinetics (PK). In our current investigation, we assessed hepatic uptake in hepatocyte suspension in Krebs-Henseleit buffer in the presence and absence of serum. The results showed that the unbound intrinsic active clearance (CLu,int,active) values obtained by normalizing the unbound fraction in the buffer containing 10% serum were generally higher than the CLu,int,active obtained directly from protein free buffer, suggesting "protein-facilitated" uptake. The differences of CLu,int,active in the buffer with and without protein ranged from 1- to 925-fold and negatively correlated to the unbound serum binding of organic anion transporting polypeptide substrates. When using the uptake values obtained from buffer containing serum versus serum-free buffer, the median of scaling factors (SFs) for CLu,int,active reduced from 24.2-4.6 to 22.7-7.1 for human and monkey, respectively, demonstrating the improvement of in vitro to in vivo extrapolation in a PBPK model. Furthermore, values of CLu,int,active were significantly higher in monkey hepatocytes than that in human, and the species differences appeared to be compound dependent. Scaling up in vitro uptake values derived in assays containing species-specific serum can compensate for the species-specific variabilities when using cynomolgus monkey as a probe animal model. Incorporating SFs calibrated in monkey and together with scaled in vitro data can be a reliable approach for the prospective human PK prediction in early drug discovery. SIGNIFICANCE STATEMENT: We investigated the protein effect on hepatic uptake in human and monkey hepatocytes and improved the in vitro to in vivo extrapolation using parameters obtained from the incubation in the present of serum protein. In addition, significantly higher active uptake clearances were observed in monkey hepatocytes than in human, and the species differences appeared to be compound dependent. The physiologically based pharmacokinetic model that incorporates scaling factors calibrated in monkey and together with scaled in vitro human data can be a reliable approach for the prospective human pharmacokinetics prediction.

Phosphorus digestibility and bioavailability in soybean meal, spray-dried plasma protein, and meat and bone meal determined using different methods.

Three experiments were conducted to determine phosphorus (P) digestibility and bioavailability using different methods. The objective of the first experiment was to determine ileal P digestibility of soybean meal (SBM), meat and bone meal (MBM), and spray-dried plasma protein (SDPP) using a precision-fed broiler chick assay. This assay involved feeding 8 g of SBM, MBM, or SDPP to broiler chicks at 21 D of age. At 6 h after feeding, ileal digesta were collected. Ileal P digestibility of SBM, MBM, and SDPP was 64, 42, and 94%, respectively. In the second experiment, ileal P digestibility and excreta P retention of SBM, SDPP, and MBM were determined using an ad libitum fed chick assay. On day 17 of age, chicks were placed on 1 of 12 dietary treatments that consisted of diets containing increasing levels of SBM, SDPP, or MBM. On day 21, ileal digesta and excreta were collected. True ileal P digestibility and true excreta P retention estimated using regression of ileal P or excreta P output on dietary P content yielded true ileal P digestibility values for SBM, SDPP, and MBM (2 diet methods for MBM) to be 83, 98, 61, and 23%, respectively. True excreta P retention values for SBM, SDPP, and MBM (2 methods) were determined to be 51, 99, 32, and 53%, respectively. The third experiment determined bioavailability of P in SBM, SDPP, and MBM relative to KH2PO4 using a chick bone ash bioassay. Dietary treatments included a P-deficient cornstarch-dextrose-SBM diet supplemented with 2 increasing levels of P from KH2PO4, SBM, SDPP, or MBM. Bioavailability of P based on tibia ash estimated using the multiple regression slope ratio method was 36, 125, and 76% for SBM, SDPP, and MBM, respectively, relative to KH2PO4. The results of this study indicated the digestibility/relative bioavailability of the P in SDPP was very high for all 3 methods, but values for SBM and MBM varied greatly among different methods.

Bilberry Supplementation after Myocardial Infarction Decreases Microvesicles in Blood and Affects Endothelial Vesiculation.

SCOPE: Diet rich in bilberries is considered cardioprotective, but the mechanisms of action are poorly understood. Cardiovascular disease is characterized by increased proatherogenic status and high levels of circulating microvesicles (MVs). In an open-label study patients with myocardial infarction receive an 8 week dietary supplementation with bilberry extract (BE). The effect of BE on patient MV levels and its influence on endothelial vesiculation in vitro is investigated. METHODS AND RESULTS: MVs are captured with acoustic trapping and platelet-derived MVs (PMVs), as well as endothelial-derived MVs (EMVs) are quantified with flow cytometry. The in vitro effect of BE on endothelial extracellular vesicle (EV) release is examined using endothelial cells and calcein staining. The mechanisms of BE influence on vesiculation pathways are studied by Western blot and qRT-PCR. Supplementation with BE decreased both PMVs and EMVs. Furthermore, BE reduced endothelial EV release, Akt phosphorylation, and vesiculation-related gene transcription. It also protects the cells from P2X7 -induced EV release and increase in vesiculation-related gene expression. CONCLUSION: BE supplementation improves the MV profile in patient blood and reduces endothelial vesiculation through several molecular mechanisms related to the P2X7 receptor. The findings provide new insight into the cardioprotective effects of bilberries.

The acid and neutral fractions of pectins isolated from ripe and overripe papayas differentially affect galectin-3 inhibition and colon cancer cell growth.

The water-soluble fractions of pectin extracted from the pulp of ripe papayas have already been found to exert positive effects on cancer cell cultures. However, the mechanisms that lead to these beneficial effects and the pectin characteristics that exert these effects are still not well understood. Characteristics such as molecular size, monosaccharide composition and structural conformation are known as polysaccharide factors that can cause alterations in cellular response. During fruit ripening, a major polysaccharide solubilization, depolymerization, and chemical modification occur. The aims of this work are to fractionate the pectin extracted from the pulp of papayas at two stages of ripening (fourth and ninth day after harvesting) into uronic and neutral fractions and to test them for the inhibition of human recombinant galectin-3 and the inhibition of colon cancer cell growth. The structures of the fractions were chemically characterized, and the uronic fraction extracted from the fourth day after harvesting presented the best biological effects across different concentrations in both galectin-3 inhibition and viability assays. The results obtained may help to establish a relationship between the chemical structures of papaya pectins and the positive in vitro biological effects, such as inhibiting cancer cell growth.

Structure-activity relationship of Citrus segment membrane RG-I pectin against Galectin-3: The galactan is not the only important factor.

Rhamnogalacturonan I (RG-I) pectin are regarded as strong galectin-3 (Gal-3) antagonist because of galactan sidechains. The present study focused on discussing the effects of more structural regions in pectin on the anti-Gal-3 activity. The water-soluble pectin (WSP) recovered from citrus canning processing water was categorized as RG-I pectin. The controlled enzymatic hydrolysis was employed to sequentially remove the α-1,5-arabinan, homogalaturonan and ß-1,4-galactan in WSP. The Gal-3-binding affinity KD (kd/ka) of WSP and debranched pectins were calculated to be 0.32 µM, 0.48 µM, 0.56 µM and 1.93 µM. Moreover, based on the more sensitive cell line (MCF-7) model, the IC30 value of WSP was lower than these of modified pectins, indicating decreased anti-Gal-3 activity. Our results suggested that the total amount of neutral sugar sidechains, the length of arabinan and cooperation between HG and RG-I played important roles in the anti-Gal-3 activity of WSP, not the Gal/Ara ratio or RG-I/HG ratio. These results provided a new insight into structure-activity relationship of citrus segment membrane RG-I as a galectin-3 antagonist and a new functional food.

Calculating safety margins using total plasma concentration versus unbound plasma concentration - does it make a difference?

Safety margin, a key aspect of any non-clinical toxicity studies, is calculated by dividing the systemic exposure (AUC) at NOAEL (No Adverse Effect Level) in toxicity studies by the clinical exposure. The validity of using total plasma concentration (Cp) to calculate AUC is often discussed, as it is the unbound plasma concentration (Cup) that elicits the pharmacological and toxicological effects. Data regarding plasma protein binding across species was collected for 114 MSD small molecule compounds which had been discontinued from development either due to non-clinical toxicity or due to clinical Adverse Effects. A >3-fold difference in unbound fraction in plasma (fup) was selected as a meaningful difference in plasma protein binding between non-clinical species and humans. In rats, dogs and non-human primates, approximately 3-5% of the compounds had a >3-fold difference in plasma protein binding than humans. Following assessment of toxicity profile of these compounds, it was concluded that calculation of safety margins after incorporating fup would have still led to the discontinuation of these compounds. Therefore, although fup can still be used for calculation of safety margin on a case by case basis, the routine use of fup for calculation of safety margins is not warranted.

Plasmonic Biosensing with Aluminum Thin Films under the Kretschmann Configuration.

Aluminum has recently attracted considerable interest as a plasmonic material due to its unique optical properties, but most work has been limited to nanostructures. We report here SPR biosensing with aluminum thin-films using the standard Kretschmann configuration that has previously been dominated by gold films. Electron-beam physical vapor deposition (EBPVD)-prepared Al films oxidize in air to form a nanofilm of Al2O3, yielding robust stability for sensing applications in buffered solutions. FDTD simulations revealed a sharp plasmonic dip in the visible range that enables measurement of both angular shift and reflection intensity change at a fixed angle. Bulk and surface tests indicated that Al films exhibited superb sensitivity performance in both categories. Compared to Au, the Al/Al2O3 layer showed a marked effect of suppressing nonspecific binding from proteins in human serum. Further characterization indicated that Al film demonstrated a higher sensitivity and a wider working range than Au films when used for SPR imaging analysis. Combined with its economic and manufacturing benefits, the Al thin-film has the potential to become a highly advantageous plasmonic substrate to meet a wide range of biosensing needs in SPR configurations.

Chip-based serum proteomics approach to reveal the potential protein markers in the sub-acute stroke patients receiving the treatment of Ginkgo Diterpene Lactone Meglumine Injection.

ETHNOPHARMACOLOGICAL RELEVANCE: Ginkgo biloba L. is a kind of traditional Chinese medicinal material with a long history. Its main active ingredients, ginkgolides, can be used for the treatment of stroke and other cardio-cerebrovascular diseases. Ginkgo Diterpene Lactone Meglumine Injection (GDLI), a modernized TCM, has attracted much attention because of its neuroprotective and anti-inflammatory properties. AIM OF THE STUDY: To uncover the effects of GDLI on ischemic stroke patients, as well as the underlying biomarkers involved in sub-acute stroke. MATERIALS AND METHODS: We used a state-of-the-art targeted proteomics chip to investigate the association between numerous serum proteins (1101 proteins) and the sub-acute phase post-ischemic stroke. Then, the relative proteins of anti-apoptosis, anticoagulant, and neuroprotection of GDLI were verified in animal models. RESULTS: Compared with the serum from healthy volunteers, we identified 15 up-regulated proteins and 26 down-regulated proteins (FC ≥ 1.5) involved in inflammatory response, immune response, and nervous system development in the sub-acute ischemic stroke. The pro-inflammatory proteins, such as IL17, MSP-R, G-CSF-R, TLR3, MIP-3ß, TNFRSF19, and TNFRSF12, were significantly increased in serum, illustrating that the chronic inflammatory state was evident in the sub-acute stage of ischemic stroke. However, the common pro-inflammatory proteins, such as IL-1ß, IL-6, IL-8, TNF-α, IFN-γ, and IL-10, known to be up-regulated in acute stroke, had close or lightly lower levels than healthy humans (FC ≥ 1.5, P > 0.05). And some cytokines (IL3, CCL13, TNFRSF3, IL10 R beta, HLA-A, IL-1 F8/FIL1 eta, TNFRSF8, CCL18) were also markedly down-regulated in the sub-acute phase of stroke. These proteins are highly associated with the onset of stroke-induced immunosuppression and post-stroke infection. Moreover, we noticed that Ginkgo Diterpene Lactone Meglumine Injection (GDLI) treatment for 14 days was helpful to the recovery of patients in the subacute period. After the treatment of GDLI, it was observed that several inflammatory cytokines (i.e. IL-17 and IL-28A), chemokine (i.e. CCL14), and Coagulation Factor III were reduced. Meanwhile, the anti-inflammatory cytokines (IL-10 R alpha, GREMLIN, and Activin C) and neurotrophic factors (Neurturin and IGFBP2) were found to be up-regulated in stroke patients through self-control observation. Finally, we identified the IGFBP2 as a novel marker in the animal models. CONCLUSIONS: In summary, the potential markers in sub-acute stroke patients were highly different from known protein markers in the acute phase of ischemic stroke. The serum protein IGFBP2 could be novel biomarkers for the treatment of GDLI in sub-acute stroke patients. Our present findings provide an innovative insight into the novel treatment of GDLI in ischemic stroke therapy.

Effects of dietary supplementation of chitosan on immune function in growing Huoyan geese.

This present experiment was performed to investigate the effects of dietary supplementation of chitosan (CS) on immune function in growing Huoyan geese. A total of 320 28-day-old healthy growing Huoyan geese (sex balance) with similar body weight were randomly allotted into control, CS100, CS200, and CS400 groups. Each group includes 4 replicates with 20 geese per replicate, and the feeding trial lasted for 4 wk. The 4 diets contained 0, 100, 200, and 400 mg CS per kg feed, respectively. The results showed that compared with the control group, the relative weight of thymus, serum concentrations of IGF-I, INS, GH, T3, T4, IgM, IgG, IgA, complement C3, and IL-2 in CS200 group were significantly higher at both 42 and 56 D of age, respectively (P < 0.05). In addition, relative weight of bursa of fabricius (BF), spleen, serum complement C4, and TNF-a concentrations in CS200 group were higher at 56 D of age (P < 0.05), no differences were observed at 42 D of age (P > 0.05). These results indicated that addition of 200 mg/kg CS enhanced immune organs weight, serum concentrations of immunoglobulins, complements, hormone, as well as cytokines, and improved immune function of growing Huoyan geese.

Discovery of a NAPE-PLD inhibitor that modulates emotional behavior in mice.

N-acylethanolamines (NAEs), which include the endocannabinoid anandamide, represent an important family of signaling lipids in the brain. The lack of chemical probes that modulate NAE biosynthesis in living systems hamper the understanding of the biological role of these lipids. Using a high-throughput screen, chemical proteomics and targeted lipidomics, we report here the discovery and characterization of LEI-401 as a CNS-active N-acylphosphatidylethanolamine phospholipase D (NAPE-PLD) inhibitor. LEI-401 reduced NAE levels in neuroblastoma cells and in the brain of freely moving mice, but not in NAPE-PLD KO cells and mice, respectively. LEI-401 activated the hypothalamus-pituitary-adrenal axis and impaired fear extinction, thereby emulating the effect of a cannabinoid CB1 receptor antagonist, which could be reversed by a fatty acid amide hydrolase inhibitor. Our findings highlight the distinctive role of NAPE-PLD in NAE biosynthesis in the brain and suggest the presence of an endogenous NAE tone controlling emotional behavior.