Succinate Promotes Phagocytosis of Monocytes/Macrophages in Teleost Fish.

Development of immunity-based strategy to manage bacterial infection is urgently needed in aquaculture due to the widespread of antibiotic-resistant bacteria. Phagocytosis serves as the first line defense in innate immunity that engulfs bacteria and restricts their proliferations and invasions. However, the mechanism underlying the regulation of phagocytosis is not fully elucidated and the way to boost phagocytosis is not yet explored. In this manuscript, we profiled the metabolomes of monocytes/macrophages isolated from Nile tilapia, prior and after phagocytosis on Vibrio alginolyticus. Monocytes/macrophages showed a metabolic shift following phagocytosis. Interestingly, succinate was accumulated after phagocytosis and was identified as a crucial biomarker to distinguish before and after phagocytosis. Exogenous succinate increased the phagocytotic rate of monocytes/macrophages in a dose-dependent manner. This effect was dependent on the TCA cycle as the inhibitor of malonate that targets succinate dehydrogenase abrogated the effect. Meanwhile, exogenous succinate regulated the expression of genes associated with innate immune and phagocytosis. In addition, succinate-potentiated phagocytosis was applicable to both gram-negative and -positive cells, including V. alginolyticus, Edwardsiella tarda, Streptococcus agalactiae, and Streptococcus iniae. Our study shed light on the understanding of how modulation on host's metabolism regulates immune response, and this can be a potent therapeutic approach to control bacterial infections in aquaculture.

Transport properties of puerarin and effect of extract of Radix Angelicae dahuricae on puerarin intestinal absorption using in situ and in vitro models.

The root of Angelica dahurica (Radix Angelicae Dahuricae, RAD), which contains coumarins and volatile oil as its main classes of active components, is often given in conjunction with Pueraria root (Radix Puerariae, RP), which contains the phytoestrogen puerarin. The two herbs are considered to be compatible 'herb-pairs' in traditional Chinese medicine. The present investigation investigates the absorption of puerarin from RP and the effect of the total coumarins and volatile oil from RAD on its absorption. The everted gut sac and single-pass intestinal perfusion methods were used, respectively. The results showed that the absorption of puerarin in the jejunum was significantly increased in the presence of the coumarins and/or volatile oil. The absorption rate constant (K(a)) of puerarin increased gradually until the concentration reached 160 µg · mL(-1), after which its absorption became saturated and the apparent permeability (P(app)) values significantly decreased. The results showed that the intestinal absorption mechanisms of puerarin involved active transportation processes and that puerarin is likely to be a substrate of P-gp because verapamil significantly affected its P(app) and K(a). The absorption of puerarin significantly increased (p < 0.01) when combined with RAD extracts, as shown by the increase in concentration of puerarin in blood from the hepatic portal vein, supporting the concept of RAD and RP as a compatible herb-pair.

[Study on transport mechanism of baicalin in Scutellariae radix extracts and effect of Angelica dahurica extracts on transport of baicalin by Caco-2 cell monolayer model].

OBJECTIVE: To study the transport mechanism of baicalin of Scutellariae Radix extracts and the effect of Angelica dahurica extracts on the intestinal absorption of baicalin by using Caco-2 cell monolayer model, in order to analyze the effect mechanism of Angelica dahurica extracts on the intestinal absorption of baicalin. METHOD: The Caco-2 cell monolayer model was established with human colonic adenocarcinoma cells, and used to study the effect of pH, time, drug concentration and temperature on the transport of baicalin in Scutellariae Radix extracts, the effect of P-gp and MRP protein-dedicated inhibitors on the bidirectional transport of baicalin in Caco-2 cell model, and the effect of angelica root extracts on baicalin absorption and transport. RESULT: Baicalin was absorbed well at 37 degrees C and under pH 7.4 condition and concentration dependent. Its proteins became inactive at 4 degrees C, with a low transport. The bi-drectional transfer PDR was 0. 54. After P-gp inhibitor verapamil and MRP inhibitor probenecid were added, the value of PappBL-AP of baicalin decreased, but without any difference in PDR. The transport of baicalin was improved by 2.34, 3.31 and 3.13 times, after A. dahurica extract coumarin, volatile oil, and mixture of coumarin and volatile oil. CONCLUSION: The transport mechanism of baicalin is mainly passive transfer and supplemented with efflux proteins involved. A. dahurica extracts can enhance the absorption of baicalin, which may be related to the passive transfer merchanism of baicalin. A. dahurica extracts' effect in opening the close junction among cells may be related to its expression or function in inhibiting efflux proteins.

Elucidation of the transport mechanism of baicalin and the influence of a Radix Angelicae Dahuricae extract on the absorption of baicalin in a Caco-2 cell monolayer model.

ETHNOPHARMACOLOGICAL RELEVANCE: Angelicae Dahurica (Hoffm.) Benth. & Hook. f. ex Franch. & Sav combined with Radix Scutellariae baicalensis Georgi has been widely used in traditional Chinese medicine (TCM) as an antipyretic analgesic and anti-inflammatory drug. Modern pharmacological studies have demonstrated that the compatible application of these two drugs is an effective treatment for hepatitis. A previous study indicated that a Radix Angelicae Dahuricae extract enhanced the intestinal absorption of the baicalin found in Radix Scutellariae; however, the underlying compatibility mechanism of these two herbs remains unknown. In this study, we further examined the effect of a Radix Angelicae Dahuricae extract on the absorption and transport properties of baicalin in a Caco-2 cell model to determine the compatibility mechanism of these two herbs. AIM OF THE STUDY: The aim of this work was to study the transport properties of baicalin in Radix Scutellariae across cell membranes and the effects of a Radix Angelicae Dahuricae extract on baicalin absorption using the well-characterized, human-based intestinal Caco-2 cell model. MATERIALS AND METHODS: We assessed the absorption, bidirectional transport and toxicity of baicalin using a range of parameters, including drug concentration, pH, a P-glycoprotein (P-gp) inhibitor (Verapamil), an MRP inhibitor (MK-571) and EDTA-Na2 (tight junction modulator). Next, we studied the influence of a Radix Angelicae Dahuricae extract on the transport of baicalin under the same conditions. Drug concentration was measured by HPLC, and the apparent permeability coefficient (Papp) and apparent permeability ratio (PDR) were subsequently calculated. RESULTS: The results showed that baicalin is non-toxic within a concentration range of 800 µg/mL to 4800 µg/mL. The transport of baicalin showed time and concentration dependence. The absorption of baicalin was optimal at pH 7.4 in 37 °C; however, the absorption decreased at 4 °C. The Papp of baicalin transport through the Caco-2 cell monolayer model was altered when specific inhibitors of P-gp or MRP were added to the cells. However, there was no significant difference in the PDR value. The Papp of baicalin improved when it was combined with the Radix Angelicae Dahuricae extract. The influence of EDTA-Na2 on the transport of baicalin showed that the permeability of baicalin significantly increased. The result further indicated that the mechanism of baicalin intestinal absorption in the Caco-2 cell monolayer involves passive transcellular diffusion. CONCLUSIONS: Passive diffusion is the main mode of intestinal absorption of bacalin and it involved in the efflux of proteins. The enhanced intestinal absorption of baicalin by Radix Angelicae Dahuricae can be due to opening of the tight junctions between cells and inhibition of MRP efflux protein expression or function.

The absorption characterization effects and mechanism of Radix Angelicae dahuricae extracts on baicalin in Radix Scutellariae using in vivo and in vitro absorption models.

ETHNOPHARMACOLOGICAL RELEVANCE: Angelicae Dahurica(Hoffm.)Benth.&Hook.f.ex Franch.&Sav combined with Scutellaria baicalensis Georgi. has been widely used as herb-pairs in traditional Chinese medicine (TCM) to treat migraine headache and cataract, but the underlying compatibility mechanism of the two herbs remains unknown. AIM OF STUDY: In the present work, we investigated the additive or synergistic effects of absorption behavior of Radix Angelicae dahuricae extracts on baicalin, and the absorption-enhancing mechanism of Radix Angelicae dahuricae extracts on baicalin. MATERIALS AND METHODS: Total coumarins (Cou) and volatile oil (VO), as the two main components of Radix Angelicae dahuricae, were extracted by supercritical fluid extraction (SFE) further treated with liquid-liquid separation method. The absorption behavior was investigated by applying the everted gut sac technique and in situ single-pass intestinal perfusion method. RESULTS AND CONCLUSIONS: The results showed that both the Cou and the VO could improve the intestinal absorption of baicalin in vivo, and had synergistic action the enhanced absorption of baicalin. Since verapamil did not affect the P(app) and K(a) of baicalin significantly, we concluded that the absorption of Baicalin could not be an active transportation in dependent of P-glycoprotein-Mediated efflux systems. Based on intestinal absorption of drug studying was one of the efficacious methods to clarify the compatibility of principles of herb-pairs. The everted gut sac technique and in situ single-pass intestinal perfusion technique model were the effective methods to study the absorption of drug, the application of the animal model to investigating the absorption of herb-drug interactions or other relevant research purposes is envisioned.

[The enhancing effect of Angelica dahurica extracts on absorption of baicalin--the active composition of Scutellaria].

To explore the mechanism of the absorption enhancement of Angelica dahurica extract (Ade), the absorption mechanism of baicalin in the Scutcllaria water extraction as well as the effect of Angelica dahurica extract on absorption of baicalin were investigated. In order to determine the main absorption site, everted intestinal sac model was used to study the effect of Angelica dahurica extract on the absorption of baicalin at duodenum, jejunum, ileum and colon. In situ single pass intestinal perfusion model was performed to study the absorption of various concentrations of baicalin and the effect of Angelica dahurica extract on the absorption of baicalin at the main absorption site. To authenticate the consequence of perfusion by getting the blood from the hepatic portal vein and determine the concentration of the baicalin in the blood. The result showed that baicalin could be absorbed at all of the four intestinal segments with increasing absorption amount per unit as follows: ileum > colon > jejunum > duodenum. The absorption ofbaicalin in the duodenum significantly increased with Angelica dahurica extract, thus, duodenum was chosen to be the studying site. Apparent permeability values (Papp) and absorption rate constant (Ka) of baicalin in the duodenum increased gradually with higher concentrations. When the concentration of baicalin rises to a certain degree, the absorption increase had a saturable process, the absorption of baicalin may be an active transportation. Baicalin may be not a substrate of P-gp as verapamil which had not significantly affected the Papp and Ka of baicalin. The absorption of baicalin in the duodenum significantly increased (P < 0.01) in the two models with Angelica dahurica extract and the concentration of baicalin in the blood from the hepatic portal vein showed that the Angelica dahurica extract can increase the absorption of baicalin.