Antifungal drug resistance is a significant clinical problem, and antifungal agents that can evade resistance are urgently needed. In infective niches, resistant organisms often co-existed with sensitive ones, or a subpopulation of antibiotic-susceptible organisms may evolve into resistant ones during antibiotic treatment and eventually dominate the whole population. In this study, we established a co-culture assay in which an azole-resistant Candida albicans strain was mixed with a susceptible strain labeled with green fluorescent protein to mimic in vivo conditions and screen for antifungal drugs. Fluconazole was used as a positive control to verify the validity of this co-culture assay. Five natural molecules exhibited antifungal activity against both susceptible and resistant C. albicans. Two of these compounds, retigeric acid B (RAB) and riccardin D (RD), preferentially inhibited C. albicans strains in which the efflux pump MDR1 was activated. This selectivity was attributed to greater intracellular accumulation of the drugs in the resistant strains. Changes in sterol and lipid compositions were observed in the resistant strains compared to the susceptible strain, and might increase cell permeability to RAB and RD. In addition, RAB and RD interfered with the sterol pathway, further aggregating the decrease in ergosterol in the sterol synthesis pathway in the MDR1-activated strains. Our findings here provide an alternative for combating resistant pathogenic fungi.
ATP-Binding Cassette Transporters ; genetics ; metabolism ; Antifungal Agents ; chemistry ; metabolism ; pharmacology ; Azoles ; pharmacology ; Biosynthetic Pathways ; drug effects ; genetics ; Candida albicans ; chemistry ; drug effects ; metabolism ; Cell Membrane ; chemistry ; metabolism ; Coculture Techniques ; Drug Resistance, Fungal ; drug effects ; Ergosterol ; metabolism ; Fungal Proteins ; genetics ; metabolism ; Lipids ; chemistry ; Molecular Structure ; Permeability ; Phenyl Ethers ; chemistry ; metabolism ; pharmacology ; Sterols ; chemistry ; metabolism ; Stilbenes ; chemistry ; metabolism ; pharmacology ; Triterpenes ; chemistry ; metabolism ; pharmacology

Single-pass intestinal perfusion( SPIP) is the common carrier of biopharmaceutics classification system( BCS) to study compound permeability. With the application and deepening study of BCS in the field of traditional Chinese medicine( TCM),SPIP model is becoming more and more common to study the intestinal absorption of TCM ingredients. Based on the limitations of the SPIP model in some researches on TCM permeability,it was speculated in this study that aglycone may be more suitable than the glycoside to study the intestinal absorption problem by using SPIP model. Furthermore,applicability of aglycone components was analyzed and evaluated. In this study,with quercetin,daidzein,formononetin,genistein and glycyrrhetinic acid used as research objects,the quantitative study of SPIP was used to evaluate the intestinal permeability of these aglycones and to predict the effective permeability coefficient( Peff) and absorption fraction( Fa) in human body. By combining studies comparison and analysis on multiple permeability research methods and prediction of human body absorption of aglycones in physiological-based pharmacokinetic models,this paper can further illustrate that the SPIP model is a good tool for studying the permeability of aglycones and predicting human absorption,which can provide data foundation and theoretical reference for researches on SPIP technique and BCS in intestinal absorption of TCM ingredients.
Biopharmaceutics ; Humans ; Intestinal Absorption ; Intestines ; drug effects ; Medicine, Chinese Traditional ; Perfusion ; Permeability

ObjectiveTo investigate the effects of Zhibai Dihuang Decoction (ZDD) on sperm mitochondrial permeability transition (MPT) in the rat model of ureaplasma urealyticum (UU) infection (UUI).

METHODSNinety male SD rats were randomly divide into five groups: normal control, UUI model control, ZDD, doxycycline, and ZDD + doxycycline. The UUI model was established in the latter four groups of rats by UU injection into the bladder. On the second day after modeling, the animals of the normal control and UUI model control groups were treated intragastrically with 0.9% sodium chloride solution and those in the other groups with corresponding drugs, all for 21 consecutive days. At 24 hours after drug withdrawal, epididymal samples were obtained for detection of the protein and mRNA expressions of VDAC2 and ANT4 in the sperm mitochondria by RT-PCR and Western blot respectively and determination of the contents of adenosine monophosphate (AMP), adenosine diphosphate (ADP) and adenosine monophosphate (AMP) and energy charge (EC) in the sperm mitochondria by high-performance liquid chromatography.

RESULTSThe protein expressions of VDAC2 and ANT4 in the rat sperm mitochondria were 0.626 ± 0.074 and 0.527 ± 0.096 in the normal control group, 0.039 ± 0.011 and 0.044 ± 0.011 in the UUI model control group, 0.101 ± 0.037 and 0.127 ± 0.040 in the ZDD group, 0.236 ± 0.070 and 0.253 ± 0.054 in the doxycycline group, and 0.475 ± 0.064 and 0.367 ± 0.086 in the ZDD + doxycycline group, significantly lower in the UUI model control than in the normal control group (P<0.05 and P<0.01), but remarkably higher in the doxycycline and ZDD + doxycycline groups than in the UUI model control (P<0.01) and the ZDD group (P<0.05 and P<0.01), and the expression of VDAC2 was markedly higher in the ZDD + doxycycline than in the doxycycline group (P<0.01). The mRNA expressions of VDAC2 and ANT4 were 0.008 ± 0.001 035 and 0.026 50 ± 0.003 401 in the normal control group, 0.000 79 ± 0.000 226 and 0.001 64 ± 0.000 205 in the UUI model controls, 0.002 06 ± 0.000 861 and 0.005 04 ± 0.002 537 in the ZDD group, 0.003 34 ± 0.000 229 and 0.008 57 ± 0. 000 690 in the doxycycline group, and 0.004 85 ± 0.000 495 and 0.013 13 ± 0.000 826 in the ZDD + doxycycline group, significantly lower in the UUI model control than in the normal control group (P<0.05 and P<0.01), but remarkably higher in the ZDD, doxycycline and ZDD + doxycycline groups than in the UUI model controls (P<0.01) as well as in the doxycycline and ZDD + doxycycline groups than in the ZDD group (P<0.01) and in the ZDD + doxycycline than in the doxycycline group (P<0.01). The levels of ATP, ADP, AMP and EC in the sperm mitochondria were (203.41 ± 13.16) mg/L, (129.87 ± 14.68) mg/L, (149.05 ± 5.65) mg/L and 0.56 ± 0.01 in the normal control group, (96.22 ± 12.55) mg/L, (99.87 ± 3.28) mg/L, (212.53 ± 19. 43) mg/L and 0.36 ± 0.03 in the UUI model control group, (101.99 ± 5.97) mg/L, (104.99 ± 16.40) mg/L, (183.97 ± 12.43) mg/L and 0.40 ± 0.01 in the ZDD group, (159.44 ± 33.16) mg/L, (118.51 ± 12.99) mg/L, (160.64 ± 14.19) mg/L and 0.50 ± 0.06 in the doxycycline group, and (194.07 ± 9.36) mg/L, (121.62 ± 9.41) mg/L, (150.21 ± 12.87) mg/L and 0.55 ± 0.01 in the ZDD + doxycycline group. The levels of ATP and EC were significantly lower and that of AMP higher in the UUI model control than in the normal control group (P<0.01), while the former two were remarkably higher and the latter one lower in the doxycycline and ZDD + doxycycline groups than in the UUI model controls (P<0.05 and P<0.01). Compared with the ZDD + doxycycline group, the ZDD group showed significantly decreased ATP and EC but increased AMP, while the doxycycline group exhibited decreases in both ATP and EC (P<0.05 and P<0.01).

CONCLUSIONSZDD can upregulate the decreased protein and mRNA expressions of VDAC2 and ANT4 in the sperm mitochondria and improve sperm mitochondrial permeability transition and mitochondrial energy metabolism in rats with UU infection, which may be one of its action mechanisms in the treatment of UU infection-induced male infertility.

Animals ; Anti-Bacterial Agents ; therapeutic use ; Doxycycline ; therapeutic use ; Drugs, Chinese Herbal ; metabolism ; therapeutic use ; Energy Metabolism ; Epididymis ; Humans ; Infertility, Male ; Male ; Mitochondria ; drug effects ; Permeability ; drug effects ; Random Allocation ; Rats ; Rats, Sprague-Dawley ; Spermatozoa ; drug effects ; Ureaplasma Infections ; drug therapy ; Ureaplasma urealyticum ; Voltage-Dependent Anion Channel 2 ; metabolism

Although a variety of regimens are available for the treatment of atopic dermatitis (AD), severe adverse reactions and unpopular costs often limit their usage. In contrast, certain inexpensive, naturally-occurring ingredients are proven effective for AD with fewer side effects. The beneficial effects of these ingredients can be attributed to inhibition of cytokine and chemokine expression, IgE production, inflammatory cell infiltration, histamine release, and/or the enhancement of epidermal permeability barrier function. Since herbal medicines are widely available, inexpensive and generally safe, they could be valuable alternatives for the treatment of AD, particularly for those patients who are not suitable for the utilization of immune modulators. In this review, we summarize the therapeutic benefits of natural ingredients for the treatment of AD and the mechanisms of their actions.
Anti-Inflammatory Agents ; pharmacology ; therapeutic use ; Biological Products ; adverse effects ; therapeutic use ; Dermatitis, Atopic ; drug therapy ; Humans ; Permeability ; Treatment Outcome

OBJECTIVETo investigate the effect of c-Met inhibitor cabozantinib (XL-184) in inhibiting Listeria monocytogenes (LM) from invading Caco-2 cells to reduce the cell injury.

METHODSThe cell invasion capacity of LM was assayed in Caco-2 cells incubated with different doses of XL-184 for different durations. Caco-2 cells incubated with XL-184 were seeded on the upper room of the transwell chamber, and the cell monolayer was exposed to LM infection followed by addition of horseradish peroxidase (HRP). The trans-epithelial electric resistance (TEER), HRP concentration and LM colony-forming unit (CFU) were measured in the cell monolayer. Fluorescent staining was used to evaluate the cell viability, and LDH release from the cells was examined to assess the changes in cell membrane permeability.

RESULTSXL-184 significantly decreased LM invasion rate in Caco-2 cells in a dose- and time-dependent manner (P=0.000), and this effect was enhanced by co-incubation of the cells with ampicillin (P<0.05). In the cell membrane permeability assay in the monolayer cells, XL-184 markedly inhibited LM-induced reduction of TEER (P<0.05) and significantly suppressed LM-induced enhancement of cell membrane permeability shown by reduced HRP concentration and LM count in the lower chamber (P=0.000). The cells infected with LM showed significantly lowered cell viability, which was rescued by XL-184 (P<0.01); XL-184 also dose-dependently reduced LDH release from the cells (P<0.05).

CONCLUSIONSXL-184 can suppress LM invasion in Caco-2 cells to reduce the cell injury, suggesting its value as a promising candidate agent for prevention and treatment of LM infections.

Anilides ; pharmacology ; Caco-2 Cells ; Cell Membrane Permeability ; drug effects ; Cell Survival ; Humans ; Listeria monocytogenes ; drug effects ; Pyridines ; pharmacology

BACKGROUNDIntact endothelial structure and function are critical for maintaining microcirculatory homeostasis. Dysfunction of the latter is an underlying cause of various organ pathologies. In a previous study, we showed that rhubarb, a traditional Chinese medicine, protected intestinal mucosal microvascular endothelial cells in rats with metastasizing septicemia. In this study, we investigated the effects and mechanisms of rhubarb on matrix metalloproteinase-9 (MMP9)-induced vascular endothelial (VE) permeability.

METHODSRhubarb monomers were extracted and purified by a series of chromatography approaches. The identity of these monomers was analyzed by hydrogen-1 nuclear magnetic resonance (NMR), carbon-13 NMR, and distortionless enhancement by polarization transfer magnetic resonance spectroscopy. We established a human umbilical vein endothelial cell (HUVEC) monolayer on a Transwell insert. We measured the HUVEC permeability, proliferation, and the secretion of VE-cadherin into culture medium using fluorescein isothiocyanate-dextran assay, 3-(4,5-dimethyl-2-thiazolyl)-2,5-diphenyl-2-H-tetrazolium bromide assay, and enzyme-linked immunosorbent assay, respectively, in response to treatment with MMP9 and/or rhubarb monomers.

RESULTSA total of 21 rhubarb monomers were extracted and identified. MMP9 significantly increased the permeability of the HUVEC monolayer, which was significantly reduced by five individual rhubarb monomer (emodin, 3,8-dihydroxy-1-methyl-anthraquinone-2-carboxylic acid, 1-O-caffeoyl-2-(4-hydroxyl-O-cinnamoyl)-β-D-glucose, daucosterol linoleate, and rhein) or a combination of all five monomers (1 μmol/L for each monomer). Mechanistically, the five-monomer mixture at 1 μmol/L promoted HUVEC proliferation. In addition, MMP9 stimulated the secretion of VE-cadherin into the culture medium, which was significantly inhibited by the five-monomer mixture.

CONCLUSIONSThe rhubarb mixture of emodin, 3,8-dihydroxy-1-methyl-anthraquinone-2-carboxylic acid, 1-O-caffeoyl-2-(4-hydroxyl-O-cinnamoyl)-β-D-glucose, daucosterol linoleate, and rhein, at a low concentration, antagonized the MMP9-induced HUVEC monolayer permeability by promoting HUVEC proliferation and reducing extracellular VE-cadherin concentrations.

Cadherins ; metabolism ; Capillary Permeability ; drug effects ; Cell Line ; Cell Proliferation ; drug effects ; Drugs, Chinese Herbal ; chemistry ; pharmacology ; Human Umbilical Vein Endothelial Cells ; cytology ; drug effects ; Humans ; Matrix Metalloproteinase 9 ; metabolism ; Rheum ; chemistry

OBJECTIVETo observe the effect of Modified Yiqi Chutan Recipe (MYCR) on blood flow perfusion in treating mid-late stage non-small cell lung cancer (NSCLC) patients by using multislice CT perfusion (CTP) , and to assess the relationship between each CTP parameter and the prognosis as well.

METHODSTotally 87 mid-late stage NSCLC patients were randomly assigned to the treatment group (44 cases, Shenyi Capsule + MYCR +chemotherapy) and the control group (43 cases, chemotherapy alone) in the ratio of 1:1. And 21 days consisted of 1 therapeutic course, 4 courses in total. All of them underwent CTP of primary tumor and routine thoracic CT examination (plain CT and enhancement CT) 3 times (before therapy, after 2 and 4 cycles). CT findings were analyzed for tumor size and perfusion parameters [blood flow (BF), blood volume (BV), permeability surface (PS), mean transit time (MTT), and time to peak (TP) before and after treatment, and relationship between perfusion parameters and prognosis was also assessed.

RESULTSIn 87 cases, 7 dropped out and 80 cases were available, 40 in the treatment group and 40 in the control group. (1) The relief rate was 47.5% (19/40) and the total stable rate was 77.5% (31/40) in the treatment group, and they were 40.0% (16/40) and 65.0% (26/40) in the control group, with no statistical difference between the two groups (χ² = 0.672, 1.227; P > 0.05). (2) Compared with before treatment group in the same group, BF and PS decreased, and MTT increased in the two groups after 2 and 4 courses (P < 0.05); BE and PS decreased, and MTT increased in the control group after 2 courses (P < 0.05). Compared with the control group after 4 courses, BE decreased more significantly in the treatment group (P < 0.05). (3) After 4 courses, all patients were assigned to the remission group (35 cases) and the non-remission group (45 cases) according to the RECIST standard. Compared with before treatment in the same group, BF, BF, and PS all decreased, and MTT increased in the remission group after treatment (all P < 0.05); BF increased in the non-remission group after treatment (P < 0.05). (4) All patients were assigned to the BE increase group (34 cases) and the BE decrease group (46 cases) according to changed BE values after treatment. Results showed the mean survival rate was 246 days in the BF increase group (the 1-year accumulative survival rate being 13.0%) and 387 days in the BE decrease group (the 1-year accumulative survival rate being 53.1%). The life span was prolonged and the 1-year accumulative survival rate was elevated in the BE increase group, with statistical difference as compared with the BE decrease group (χ² = 19.057, P < 0.01).

CONCLUSIONSShenyi Capsule plus MYCR could reduce BE in mid-late stage NSCLC patients , improve vascular permeability, showing better synergistic effect with chemotherapy. CTP could not only reflect the change of tumor size, but also reflect vascular function of the tumor. Meanwhile, changes of CTP parameters were closely associated with prognosis. Patients with post-treatment decreased BE value had better prognosis and longer life span.

Capillary Permeability ; drug effects ; Carcinoma, Non-Small-Cell Lung ; diagnosis ; drug therapy ; Drugs, Chinese Herbal ; therapeutic use ; Humans ; Phytotherapy ; Tomography, X-Ray Computed

OBJECTIVETo investigate the effect of epigallocatechin-3-gallate (EGCG) on biomodification of demineralized dentine substrate, in its permeability, hydrophobicity, and inhibition ability to collagen enzymatic degradation.

METHODSThe dentine substrates were treated with simulated pulpal pressure created by mixtures of 0.02%, 0.1% EGCG/bovine serum albumin (BSA) in acidic environment (pH4.4) for 48 h. A fluid-transport model was used to measure the fluid permeability through demineralized dentine substrate. Positive replicas of dentine substrate were fabricated before and after being subjected to acidic environment for scanning electron microscope (SEM) examination. The blank group contained no EGCG and the positive group were treated with Gluma desensitizer. Static contact angle measurements on demineralized dentin and 0.1% EGCG primed dentin were performed by contact angle analyzer. The priming time were 60 s, 120 s, 0.5 h, 1 h. Dentine specimens bonded with Adper single bond 2 were subjected to 100 mg/L collagenase and observed under SEM. Resin-bonded specimens (with 0.02%, 0.1%, 0.5% EGCG priming, or without EGCG priming) were created for micro-tensile bond strength evaluation (MTBS). Resin-bonded specimens after thermol cycling were created for MTBS evaluation.

RESULTSThe fluid permeability in the blank control group increased ([151.3±22.3]%), the fluid permeability in 0.1% EGCG/BSA group decreased ([23.7±6.3]%). Compared to the blank control group, the contact angle of 120 s, 0.5 h, 1 h groups increased by 31.0%, 53.5%, 57.8% in deep dentin and 37.4%, 59.3%, 62.4% in shallow dentin. The SEM examination showed that 0.1% and 0.5% EGCG priming for 120 s significantly increased dentin collagen's resistance to collagenase. The immediate MTBS of 0.1% and 0.5% EGCG groups were (29.4±4.8) and (19.8± 4.9) MPa. After thermol cycling, the MTBS of 0.1% and 0.5% EGCG groups were (19.9±5.1) and (15.3± 6.3) MPa.

CONCLUSIONSUnder acidic environment (pH4.4), the 0.1% EGCG can reduce dentine permeability under acidic environment. The 0.1% EGCG can increase hydrophobicity of dentin substrate, and strengthen dentin substrate's resistance to collagenase hydrolysis, thus increased the resin-dentin bonding durability.

Acid Etching, Dental ; Catechin ; analogs & derivatives ; pharmacology ; Collagen ; chemistry ; drug effects ; Collagenases ; pharmacology ; Composite Resins ; Dental Bonding ; Dental Cements ; Dental Pulp ; Dentin ; chemistry ; drug effects ; Dentin Permeability ; drug effects ; Dentin-Bonding Agents ; Glutaral ; pharmacology ; Hydrogen-Ion Concentration ; Hydrolysis ; Methacrylates ; pharmacology ; Microscopy, Electron, Scanning ; Pressure ; Resin Cements ; Serum Albumin, Bovine ; pharmacology ; Tensile Strength ; Time Factors

OBJECTIVETo observe the effect of natural borneol on the permeability of blood tumor barrier (BTB) model and the expression and activation of mitogen-activated protein kinase (MAPKs) signal transduction pathway related protein kinase in vitro.

METHODSC6 rat glioma cells and human umbilical vein endothelial cells (HUVECs) were co-cultured to establish BTB model. Then 4 groups were set up, the blank control group, low, middle, and high dose borneol groups (25, 50, 100 µg/mL), 3 samples collected at 7 time points (0, 10, 30, 60, 120, 180, 240 min, respectively). Blank culture medium was exchanged in the blank control group while medication. Different doses of natural borneol were administered to the 3 borneol groups. Cells were collected at different time points. BTB permeability was determined using horseradish peroxidase (HRP). Expression levels of extracellular signal regulated protein kinase (ERK), phosphorylation extracellular signal regulated protein kinase (P-ERK), P38MAPK, phosphor-P38MAPK, c-Jun N-terminal kinase (JNK), and phosphorylation c-Jun N-terminal kinase (P-JNK) were detected using Western blot.

RESULTSCompared with the same group at min 0, the permeation rate obviously increased (P < 0.01) in the 3 borneol groups at the rest time points. P-ERK expression was elevated first, reached the peak at 30 min, and gradually recovered to the initial level (P > 0.05). Compared with the blank control group, HRP permeation rate increased from 10 min to 240 min (P < 0.01), and expression of P-ERK protein increased at 30 min and 60 min (P < 0.05) in the low dose borneol group; expression of P-JNK protein decreased in the 3 borneol groups at 180 min and 240 min (P < 0.05). Compared with the low dose borneol group, expression of P-ERK protein increased from 10 min to 180 min (P < 0.05), HRP permeation rate increased from 30 min to 180 min (P < 0.05), expression of P-JNK protein decreased at 180 and 240 min (P < 0.05) in the middle dose borneol group. Compared with the middle dose borneol group, HRP permeation rate increased from 10 min to 180 min (P < 0.05), expression of P-ERK protein increased from 10 min to 180 min (P < 0.05), expression of P-JNK protein increased at 180 min and decreased at 240 min (both P < 0.05) in the high dose borneol group.

CONCLUSIONNatural borneol arrived at the effect of regulating reversible BTB patency possibly through activating phosphorylation of ERK in MAPKs signal transduction pathway, and further reversibly down-regulating expression of associated proteins.

Animals ; Bornanes ; pharmacology ; Cell Line, Tumor ; drug effects ; Coculture Techniques ; Extracellular Signal-Regulated MAP Kinases ; metabolism ; Glioma ; pathology ; Human Umbilical Vein Endothelial Cells ; Humans ; JNK Mitogen-Activated Protein Kinases ; metabolism ; Neoplasms ; pathology ; Permeability ; Phosphorylation ; Rats ; Signal Transduction ; drug effects ; p38 Mitogen-Activated Protein Kinases ; metabolism

Pulmonary dysfunction caused by ischemia-reperfusion injury is the leading cause of mortality in lung transplantation. We aimed to investigate the effects of sevoflurane pretreatment on lung permeability, tight junction protein occludin and zona occludens 1 (ZO-1) expression, and translocation of protein kinase C (PKC)-alpha after ischemia-reperfusion. A lung ischemia-reperfusion injury model was established in 96 male Wistar rats following the modified Eppinger method. The rats were divided into four groups with 24 rats in each group: a control (group C), an ischemia-reperfusion group (IR group), a sevoflurane control group (sev-C group), and a sevoflurane ischemia-reperfusion group (sev-IR group). There were three time points in each group: ischemic occlusion for 45 min, reperfusion for 60 min and reperfusion for 120 min; and there were six rats per time point. For the 120-min reperfusion group, six extra rats underwent bronchoalveolar lavage. Mean arterial pressure (MAP) and pulse oxygen saturation (SpO2) were recorded at each time point. The wet/dry weight ratio and lung permeability index (LPI) were measured. Quantitative RT-PCR and Western blot were used to measure pulmonary occludin and ZO-1, and Western blot was used to measure cytosolic and membranous PKC-alpha in the lung. Lung permeability was significantly increased after ischemia-reperfusion. Sevoflurane pretreatment promoted pulmonary expression of occludin and ZO-1 after reperfusion and inhibited the translocation of PKC-alpha. In conclusion, sevoflurane pretreatment alleviated lung permeability by upregulating occludin and ZO-1 after ischemia-reperfusion. Sevoflurane pretreatment inhibited the translocation and activation of PKC-alpha, which also contributed to the lung-protective effect of sevoflurane.
Anesthetics, Inhalation/*therapeutic use ; Animals ; Capillary Permeability/drug effects ; Gene Expression Regulation/drug effects ; Lung/*drug effects/metabolism/pathology ; Lung Diseases/*drug therapy/genetics/metabolism/pathology ; Male ; Methyl Ethers/*therapeutic use ; Protein Kinase C-alpha/*metabolism ; Protein Transport/drug effects ; RNA, Messenger/genetics ; Rats, Wistar ; Reperfusion Injury/*drug therapy/genetics/metabolism/pathology ; Zonula Occludens-1 Protein/analysis/*genetics