Aedes albopictus salivary adenosine deaminase is an immunomodulatory factor facilitating dengue virus replication.

The Asian tiger mosquito, Aedes albopictus, is an important vector for the transmission of arboviruses such as dengue virus (DENV). Adenosine deaminase (ADA) is a well-characterized metabolic enzyme involved in facilitating blood feeding and (or) arbovirus transmission in some hematophagous insect species. We previously reported the immunologic function of ADA by investigating its effect on mast cell activation and the interaction with mast cell tryptase and chymase. The 2-D gel electrophoresis and mass spectrometry analysis in the current study revealed that ADA is present and upregulated following mosquito blood feeding, as confirmed by qRT-PCR and western blot. In addition, the recombinant ADA efficiently converted adenosine to inosine. Challenging the Raw264.7 and THP-1 cells with recombinant ADA resulted in the upregulation of IL-1ß, IL-6, TNF-α, CCL2, IFN-ß, and ISG15. The current study further identified recombinant ADA as a positive regulator in NF-κB signaling targeting TAK1. It was also found that recombinant Ae. albopictus ADA facilitates the replication of DENV-2. Compared with cells infected by DENV-2 alone, the co-incubation of recombinant ADA with DENV-2 substantially increased IL-1ß, IL-6, TNF-α, and CCL2 gene transcripts in Raw264.7 and THP-1 cells. However, the expression of IFN-ß and ISG15 were markedly downregulated in Raw264.7 cells but upregulated in THP-1 cells. These findings suggest that the immunomodulatory protein, Ae. albopictus ADA is involved in mosquito blood feeding and may modulate DENV transmission via macrophage or monocyte-driven immune response.

A C-type lectin in saliva of Aedes albopictus (Diptera: Culicidae) bind and agglutinate microorganisms with broad spectrum.

Via complex salivary mixture, mosquitos can intervene immune response and be helpful to transmit several viruses causing deadly human diseases. Some C-type lectins (CTLs) of mosquito have been reported to be pattern recognition receptor to either resist or promote pathogen invading. Here, we investigated the expression profile and agglutination function of an Aedes albopictus CTL (Aalb_CTL2) carrying a single carbohydrate-recognition domain (CRD) and WND/KPD motifs. The results showed that Aalb_CTL2 was found to be specifically expressed in mosquito saliva gland and its expression was not induced by blood-feeding. The recombinant Aalb_CTL2 (rAalb_CTL2) could agglutinate mouse erythrocytes in the presence of calcium and the agglutinating activity could be inhibited by EDTA. rAalb_CTL2 also displayed the sugar binding ability to D-mannose, D-galactose, D-glucose, and maltose. Furthermore, it was demonstrated that rAalb_CTL2 could bind and agglutinate Gram positive bacteria Staphylococcus aureus and Bacillus subtilis, Gram negative bacteria Escherichia coli and Pseudomonas aeruginosa, as well as fungus Candida albicans in vitro in a calcium dependent manner. However, rAalb_CTL2 could not promote type 2 dengue virus (DENV-2) replication in THP-1 and BHK-21 cell lines. These findings uncover that Aalb_CTL2 might be involved in the innate immunity of mosquito to resist microorganism multiplication in sugar and blood meals to help mosquito survive in the varied natural environment.

Strain differences in the drug transport capacity of intestinal glucose transporters in Sprague-Dawley versus Wistar rats, C57BL/6J versus Kunming mice.

Designing oral drug delivery systems using intestinal glucose transporters (IGTs) may be one of the strategies for improving oral bioavailability of drugs. However, little is known about the biological factors affecting the drug transport capacity of IGTs. Gastrodin is a sedative drug with a structure very similar to glucose. It is a highly water-soluble phenolic glucoside. It can hardly enter the intestine through simple diffusion but exhibits good oral bioavailability of over 80%. We confirmed that gastrodin is absorbed via the intestinal glucose transport pathway. It has the highest oral bioavailability among the reported glycosides' active ingredients through this pathway. Thus, gastrodin is the most selective drug substrate of IGTs and can be used to evaluate the drug transport capacity of IGTs. Obviously, strain is one of the main biological factors affecting drug absorption. This study firstly compared the drug transport capacity of IGTs between SD rats and Wistar rats and between C57 mice and KM mice by pharmacokinetic experiments and single-pass intestinal perfusion experiments of gastrodin. Then, the sodium-dependent glucose transporter type 1 (SGLT1) and sodium-independent glucose transporters type 2 (GLUT2) in the duodenum, jejunum, ileum and colon of these animals were quantified using RT-qPCR and Western blot. The results showed that the oral bioavailability of gastrodin in Wistar rats was significantly higher than in SD rats and significantly higher in KM mice than in C57 mice. Gastrodin absorption significantly differed among different intestinal segments in SD rats, C57 mice and KM mice, except Wistar rats. RT-qPCR and Western blot demonstrated that the intestinal expression distribution of SGLT1 and GLUT2 in SD rats and C57 mice was duodenum ≈ jejunum > ileum > colon. SGLT1 expression did not differ among different intestinal segments in KM mice, whereas the intestinal expression distribution of GLUT2 was duodenum ≈ jejunum ≈ ileum > colon. However, the expression of SGLT1 and GLUT2 did not differ among different intestinal segments in Wistar rats. It was reported that the intestinal expression distribution of SGLT1 and GLUT2 in humans is duodenum > jejunum > ileum > colon. Hence, the intestinal expression distribution of SGLT1 and GLUT2 of SD rats and C57 mice was more similar to that in humans. In conclusion, the drug transport capacity of IGTs differs in different strains of rats and mice. SD rats and C57 mice are more suitable for evaluating the pharmacokinetics of glycosides' active ingredients absorbed via the intestinal glucose transport pathway.

Preparation, Characterization, and Selection of Optimal Forms of (S)-Carvedilol Salts for the Development of Extended-Release Formulation.

(S)-carvedilol (S-CAR) is the dominant pharmacodynamic conformation of carvedilol, but its further development for extended-release formulation is restricted by its poor solubility. This study aimed to prepare and screen S-CAR salts that could be used to improve solubility and allow extended release. Five salts of S-CAR with well-known acid counterions (i.e., phosphate, hydrochloride, sulfate, fumarate, and tartrate) were produced using similar processes. However, these salts were obtained with water contents of 1.60-12.28%, and their physicochemical properties differed. The melting points of phosphate, hydrochloride, and tartrate were 1.1-1.5 times higher than that of the free base. The solubility of S-CAR salts was promoted to approximately 3-32 times higher than that of the free base at pH 5.0-8.0. Typical pH-dependent solubilities were evidently observed in S-CAR salts, but considerable differences in solubility properties among these salts were observed. S-CAR phosphate and hydrochloride possessed high melting points, considerable solubility, and excellent chemical and crystallographic stabilities. Accordingly, S-CAR phosphate and hydrochloride were chosen for further pharmacokinetic experiments and pharmaceutical study. S-CAR phosphate and hydrochloride extended-release capsules were prepared using HPMC K15 as the matrix and presented extended release in in vitro and in vivo evaluations. Results implied that water molecules in the hydrated salt were a potential threat to the achievement of crystal stability and thermostability. S-CAR phosphate and hydrochloride are suitable for further development of the extended-release formulation.

Comparative study on the intestinal absorption of three gastrodin analogues via the glucose transport pathway.

Gastrodin is the main active constituent of Tianma, a famous traditional Chinese herbal medicine. Our previous research has found that gastrodin is absorbed rapidly in the intestine by the sodium-dependent glucose transporter 1 (SGLT1). In the current report, gastrodin is the best glycoside compound absorbed via the glucose transport pathway. This study aimed to investigate the effect of the slight difference in chemical structure on the drug intestinal absorption via the glucose transport pathway. Traditional biopharmaceutical and computer-aided molecular docking methods were used to evaluate the intestinal absorption characteristics of three gastrodin analogues, namely, salicin, arbutin and 4-methoxyphenyl-ß-D-glucoside (4-MG). The oil-water partition coefficient (logP) experiments showed that the logP values of the gastrodin analogues followed the order: 4-MG > salicin > arbutin. In vitro Caco-2 cell transport experiments demonstrated that the apparent permeability coefficient (Papp) value of arbutin was higher than those of salicin and 4-MG. In situ single-pass intestinal perfusion experiments showed that the absorption of arbutin and 4-MG was better than that of salicin and that the absorption of the three compounds in the colon was lower than that in the small intestine. Quantitative real-time polymerase chain reaction results confirmed that the SGLT1 mRNA expression in the small intestine of rats was obviously higher than that in the colon of rats. In vivo pharmacokinetic experiments demonstrated that the oral bioavailability of salicin was lower than those of arbutin and 4-MG. In vitro and in vivo experiments showed that glucose or phlorizin (SGLT1 inhibitor) could decrease the intestinal absorption of the three compounds. Contrary to the above biopharmaceutical experiments, the computer-aided molecular docking test showed that the affinity of salicin to the vSGLT receptor was stronger than those of arbutin and 4-MG. In conclusion, the SGLT1 can facilitate the intestinal absorption of salicin, arbutin and 4-MG, and the slight difference in chemical structure can affect absorption.

Determination and Comparison of the Solubility, Oil-Water Partition Coefficient, Intestinal Absorption, and Biliary Excretion of Carvedilol Enantiomers.

Carvedilol is administered as a racemic mixture for the therapy of hypertension and heart failure. S-enantiomer is the dominant conformation of pharmacodynamics, but its further development was obstructed by its poor bioavailability. In this study, carvedilol and its enantiomers were compared in terms of solubility, permeability, and biliary excretion, and reasons for the poor bioavailability were discussed. Equilibrium solubility and log P were measured by a shake flask method at a wide pH range (1.2-8.0), and intestinal absorption and biliary excretion were evaluated using a single-pass rat intestinal perfusion model. According to BCS guidance, carvedilol and its R/S enantiomers are considered highly soluble at pH value less than 5.0 and low soluble at neutral or weak alkaline conditions. RS-carvedilol showed significantly lower solubilities at pH 1.2-5.0 and higher solubilities at pH 6.0-8.0 than its enantiomers. In addition, carvedilol and its enantiomers possessed similar log P values at pH 1.2-8.0. High intestinal permeabilities of carvedilol and its enantiomers were observed, and S-carvedilol showed higher absorption than R-carvedilol and RS-carvedilol. The biliary excretion about two major metabolites, 1-hydroxycarvedilol O-glucuronide and 8-hydroxycarvedilol O-glucuronide, of RS-carvedilol, S-carvedilol, and R-carvedilol were 66.4%, 73.5%, and 54.3%, respectively. In conclusion, there are significant differences amongst carvedilol and its R/S enantiomers in terms of solubility, intestine absorption, and biliary excretion abilities. The first pass effect is the primary reasons for the low bioavailability of S-carvedilol. Therefore, pharmaceutical strategies or parenteral routes should be considered to avoid the first pass metabolism.

Optimization of lipid materials in the formulation of S-carvedilol self-microemulsifying drug-delivery systems.

OBJECTIVES: The blocking effect of S-carvedilol (S-CAR) on the beta-adrenoceptor is about 100 times stronger than that of the right-handed conformation. However, further development is restricted because of its poor bioavailability caused by its low solubility and high first-pass effect. In the study, S-CAR self-microemulsifying drug-delivery systems (SMEDDSs) were established, and the effects of different lipid materials on the absorption and metabolism of S-CAR were investigated. METHODS: Six kinds of lipid materials with different chemical structures including oleic acid, glycerol monooleate, glycerol trioleate, oleoyl macrogol-6 glycerides, soybean lecithin, and α-tocopherol were selected to be the oil phase. The S-CAR SMEDDSs were prepared by the same ratio. In vitro characteristics, in vitro release, in situ intestine absorption, and bile excretion, as well as the in vivo characteristic of relative bioavailability, were determined. KEY FINDINGS: The lipid structure significantly affected physical characteristics, the absorption and excretion rates of S-CAR SMEDDSs. The findings of rat-intestine perfusion experiments showed that the S-CAR SMEDDSs decreased the bile-excretion rate of S-CAR. Compared with the S-CAR group, the oleic acid and soybean lecithin groups decreased the bile excretion to 32% and 45%, respectively. Pharmacokinetic studies showed that the AUCs of these two groups were about 1.9 and 1.7 times more than that of the S-CAR group, and the mean retention time was extended. CONCLUSION: The SMEDDS using ionic lipids (oleic acid or soybean lecithin) as oil phase can increase the oral bioavailability of S-CAR by increasing the solubility and reducing the first-pass effect.

Role of Glucose Transporters in Drug Membrane Transport.

BACKGROUND: Glucose is the main energy component of cellular activities. However, as a polar molecule, glucose cannot freely pass through the phospholipid bilayer structure of the cell membrane. Thus, glucose must rely on specific transporters in the membrane. Drugs with a similar chemical structure to glucose may also be transported through this pathway. METHODS: This review describes the structure, distribution, action mechanism and influencing factors of glucose transporters and introduces the natural drugs mediated by these transporters and drug design strategies on the basis of this pathway. RESULTS: The glucose transporters involved in glucose transport are of two major types, namely, Na+-dependent and Na+-independent transporters. Glucose transporters can help some glycoside drugs cross the biological membrane. The transmembrane potential is influenced by the chemical structure of drugs. Glucose can be used to modify drugs and improve their ability to cross biological barriers. CONCLUSION: The membrane transport mechanism of some glycoside drugs may be related to glucose transporters. Glucose modification may improve the oral bioavailability of drugs or achieve targeted drug delivery.