BAMBI regulates macrophages inducing the differentiation of Treg through the TGF-ß pathway in chronic obstructive pulmonary disease.

BACKGROUND: Chronic obstructive pulmonary disease (COPD) is characterized by continuous flow limitation and the immune system including macrophages and regulatory T lymphocytes (Tregs) is involved in COPD pathogenesis. In our previous study, we investigated that TGF-ß/BAMBI pathway was associated with COPD by regulating the balance of Th17/Treg. However, the role of bone morphogenetic protein and activin membrane-bound inhibitor (BAMBI), a pseudoreceptor of TGF-ß signalling pathway, in regulating the immune system of COPD patients has not been fully studied. Hence, we speculate that the pseudoreceptor BAMBI may play roles in the regulation of M2 macrophages to induce the differentiation of CD4+ naïve T cells into Tregs and influence the immune response in COPD. METHODS: Peripheral blood mononuclear cells (PBMCs) were isolated from healthy nonsmokers (n = 12), healthy smokers (n = 10) and COPD patients (n = 20). Naïve CD4+ T cells and monocytes-induced macrophages were used for coculture assays. The phenotypic characteristics of macrophages and Tregs were determined by flow cytometry. The expression levels of BAMBI and the TGF-ß/Smad pathway members in M2 macrophages were measured by a Western blot analysis. The monocyte-derived macrophages were stimulated with cigarette smoke extract (CSE, concentration of 0.02%) to simulate the smoking process in humans. pCMV-BAMBI was transfected into monocyte-derived M2 macrophages for subsequent co-culture assays and signalling pathway analysis. RESULTS: Our results showed that M2 macrophages could induce the differentiation of Tregs through the TGF-ß/Smad signalling pathway. In addition, monocyte-derived macrophages from COPD patients highly expressed BAMBI, and had a low capacity to induce Tregs differentiation. The expression of BAMBI and the forced expiratory volume in 1 second (FEV1%) were negatively correlated in COPD. Furthermore, overexpression of BAMBI promoted the conversion of M2 macrophages to M1 macrophages via the TGF-ß/Smad pathway. CONCLUSIONS: We demonstrated that BAMBI could promote the polarization process of M2 macrophages to M1 macrophages via the TGF-ß/Smad signalling pathway and that overexpression of BAMBI could decrease the ability of M2 macrophages to induce Treg differentiation. These findings may provide a potential mechanism by which blocking BAMBI could improve immune function to regulate COPD inflammatory conditions.

Identification and Pharmacokinetic Studies on Complanatuside and Its Major Metabolites in Rats by UHPLC-Q-TOF-MS/MS and LC-MS/MS.

The metabolic and pharmacokinetic studies on complanatuside, a quality marker of a Chinese materia medicatonic, Semen Astragali Complanati, were carried out. The UHPLC-Q-TOF/MS (ultra-high performance liquid chromatography coupled with electrospray ionization tandem quadrupole-time-of-flight mass spectrometry) method was applied to identify the metabolites of complanatuside in rat plasma, bile, stool, and urine after oral administration at the dosage of 72 mg/kg. Up to 34 metabolites (parent, 2 metabolites of the parent drug, and 31 metabolites of the degradation products) were observed, including processes of demethylation, hydroxylation, glucuronidation, sulfonation, and dehydration. The results indicated glucuronidation and sulfonation as major metabolic pathways of complanatuside in vivo. Meanwhile, a HPLC-MS method to quantify complanatuside and its two major metabolites-rhamnocitrin 3-O-ß-glc and rhamnocitrin-in rat plasma for the pharmacokinetic analysis was developed and validated. The Tmax (time to reach the maximum drug concentration) of the above three compounds were 1 h, 3 h, and 5.3 h, respectively, while the Cmax (maximum plasma concentrations)were 119.15 ng/mL, 111.64 ng/mL, and 1122.18 ng/mL, and AUC(0-t) (area under the plasma concentration-time curve) was 143.52 µg/L·h, 381.73 µg/L·h, and 6540.14 µg/L·h, accordingly. The pharmacokinetic characteristics of complanatuside and its two metabolites suggested that complanatuside rapidly metabolized in vivo, while its metabolites-rhamnocitrin-was the main existent form in rat plasma after oral administration. The results of intracorporal processes, existing forms, and pharmacokinetic characteristics of complanatuside in rats supported its low bioavailability.

Qualitative and Quantitative Analysis of the Major Constituents in WLJ Herbal Tea Using Multiple Chromatographic Techniques.

Quality control of Chinese herbal tea remains a challenge due to our poor knowledge of their complex chemical profile. This study aims to investigate the chemical composition of one of the best-selling and famous brand of beverage in China, Wanglaoji Herbal Tea (WLJHT), via a full component quantitative analysis. In this paper, a total of thirty-two representative constituents were identified or tentatively characterized using ultra-high performance liquid chromatography coupled with quadrupole tandem time-of-flight mass spectrometry (UPLC-Q-TOF-MS). Moreover, the quantitative analyses of fourteen constituents were performed by high performance liquid chromatography with a triple quadruple tandem mass spectrometry (HPLC-MS/MS) method and saccharide compositions of WLJHT were also quantitatively determined by high performance liquid chromatography (HPLC) with evaporative light scattering detector (ELSD) on a Hilic column, separately. Using multiple chromatographic techniques presented a good precision, sensitivity, repeatability and stability, and was successfully applied to analyze 16 batches of WLJHT samples. Therefore, it would be a reliable and useful approach for the quality control of WLJHT.

[Interaction of phenylethanoid glycosides and calf thymus DNA].

In physiological condition, the interaction of acteoside and forsythoside B with calf thymus DNA using neutral red (NR) as a fluorescence probe were investigated by fluorescence, UV-visible spectrophotometry, viscosity, DNA melting techniques, and molecular docking. It is observed that acteoside and forsythoside B can react with DNA. The major mode of recognition between drug and DNA is groove binding by hydrogen bonds, and the interaction of acteoside with DNA is stronger than that of forsythoside B.

[HPLC Fingerprints of Nerviliae fordii and Nervilia plicata].

OBJECTIVE: To establish the HPLC fingerprint of the whole plant of Nerviliae fordii and Nervilia plicata, in order to provide a method for evaluation of authenticity and quality control of the whole plant of Nerviliae fordii. METHODS: 15 batches of Nerviliae fordii and 5 batches of Nervilia plicata were analyzed on a Kromasil C18 column (250 mm x 4.6 mm, 5 µm) at room temperature with gradient elution using acetonitrile and 0.2% phosphoric acid as the mobile phase at a flow rate of 1.0 mL/min. The wavelength of detector was 256 nm. RESULTS: Significant differences were found between Nerviliae fordii and Nervilia plicata. Furthermore, there were no obvious differences observed between Nerviliae fordii of big-leaf and Nerviliae fordii of small-leaf. CONCLUSION: The present-developed HPLC fingerprints method provides a rapid, effective and valuable benchmark for distinguishing of Nerviliae fordii and Nervilia plicata, which is favorable to improve overall quality control of Nerviliae Fordii.

[Simultaneous determination of 4 alkaloids and a flavonoid in Picrasmae Ramulus et Folium by RP-HPLC].

A RP-HPLC method was developed to evaluate the quality of Picrasmae Ramulus et Folium by simultaneous determination of five constituents including 1-hydroxymethyl-beta-carboline (1), 1-methoxicabony-beta-carboline (2), 4-methoxy-5-hydroxy-canthin-6-one (3), 4, 5-dimethoxy-canthin-6-one (4) and maackiain (5) in Picrasmae Ramulus et Folium. The samples were separated on a Kromasil RP-C18 (4.6 mm x 250 mm, 5 microm) column eluted with acetonitrile and 0.1% phosphoric acid as mobile phases in gradient mode. The detection wavelength was set at 254 nm. The calibration curves and linearity of the above five standards were determined as (1) Y = 6 525.6X + 37.25 (0.009-1.780 microg, r = 0.996 8), (2) Y = 3 662.3X + 41.55 (0.005-0.920 microg, r = 0.999 5), (3) Y = 3763.1X + 146.87 (0.015-3.060 microg, r = 0.999 0), (4) Y = 2 174.1X + 21.52 (0.003-0.620 microg, r = 0.999 5), and (5) Y = 276.25X + 7.65 (0.010-1.960 microg, r = 0.998 9), respectively. The method is simple and repeatable, and can be used for the quality assessment of Picrasmae Ramulus et Folium.

[Study on chemical constituents of Callicarpa peii].

OBJECTIVE: To study the chemical constituents of Callicarpa peii. METHODS: The chemical constituents were isolated and purified by chromatographic methods and elucidated by spectral analysis, including UV, IR, MS, 1H-NMR and 13C-NMR. RESULTS: Ten compounds were obtained and identified as oleanolic acid (1), 2beta, 3beta,19alpha-trihydroxy-12-en-28-ursolic acid (2), luteolin -7,4'-dimethylether (3), luteolin -3', 4', 7, -trimethylether (4), luteolin -4'-methylether (5), hydnocarpin (6), luteolin (7), lyoniresinol 3alpha-O-beta-D-glucopyranoside (8), kelampayoside A (9), kaempferol -3-O-glucuronide (10). CONCLUSION: All these compounds are isolated from Callicarpa peii for the first time and compounds 3, 4, 6, 8 and 10 are isolated from this genus for the first time.

Improving ocular bioavailability of hydrophilic drugs through dynamic covalent complexation.

The clinical benefits of diquafosol tetrasodium (DQS), a hydrophilic P2Y2 receptor agonist for dry eye, have been hindered by a demanding dosing regimen. Nevertheless, it is challenging to achieve sustained release of DQS with conventional drug delivery vehicles which are mainly designed for hydrophobic small molecule drugs. To address this, we developed an affinity hydrogel for DQS by taking advantage of borate-mediated dynamic covalent complexation between DQS and hydroxypropyl guar. The resultant formulation (3% DQS Gel) was characterized by sustained release, low corneal permeation, and extended ocular retention, which were desirable attributes for ocular surface drug delivery. Both in vitro and in vivo studies had been carried out to verify the biocompatibility of 3% DQS Gel. Using corneal fluorescein staining, the Schirmer's test, PAS staining, quantitative PCR and immunohistological analyses as outcome measures, the superior therapeutic effects of 3% DQS Gel over PBS, the hydrogel vehicle and free DQS were demonstrated in a mouse dry eye model. Our DQS delivery strategy reported herein is readily applicable to other hydrophilic small molecule drugs with cis-diol moieties, thus providing a general solution to improve clinical outcomes of numerous diseases.

Triterpenes from Callicarpa integerrima Champ.

A new triterpenoid saponin and fourteen known triterpenoids were isolated from the methanol extract of the stems and leaves of Callicarpa integerrima Champ, which is used in Chinese folk medicine for stopping bleeding, expelling the wind, dissipating stagnation, and treating scrofula, by using various chromatographies, such as silica gel, Sephadex LH-20 and RP-C18 column chromatography. Their structures were identified as a new compound 2alpha, 3beta, 19alpha, 23-tetrahydroxy-olean-12-en-28-oic acid-28-O-beta-D-glucopyranosyl-(1 --> 4)-beta-D-glucopyranoside (1), together with fourteen known compounds: oleanolic acid (2), 3-acetyl oleanolic acid (3), 3beta-O-acetyl ursolic acid (4), 2alpha-hydroxy-ursolic acid (5), 2alpha, 3beta, 19alpha, 23-tetrahydroxy-urs-12-en-28-oic acid (6), alpha-amyrin-3-O-beta-D-glucopyranoside (7), pomolic acid (8), betulinic acid (9), ursolic acid (10), 2alpha, 3beta, 19alpha, 23-tetrahydroxy-olean-12-en-28-oic acid (arjungenin) (11), 2alpha-hydroxy-oleanolic acid (12), hederagenin (13), 2alpha, 19alpha-dihydroxy-ursolic acid (14) and pruvuloside A (15), by the spectroscopic techniques of NMR, HMBC, IR and MS, separately. All these compounds were obtained from this plant for the first time, and compounds 3, 4 and 15 were isolated from genus Callicarpa L. for the first time.

Enhanced analytical performance of disposable 3D carbon electrodes prepared with stainless steel wire mesh.

This paper aims to use low-cost stainless steel wire mesh (SSWM) as uniform templates to prepare disposable three-dimensional (3D) carbon electrodes to improve their analytical performance. Native SSWM electrodes were prepared with lamination and then coated with carbon cement for bulk preparation of disposable 3D carbon electrodes with drop-casting. The electrodes were then coupled in paper-based analytical devices. Meanwhile, disposable 2D carbon electrodes were prepared with the stainless steel sheets (SSSs) for comparison under the same condition using stripping analysis of heavy metals as a model. Our results demonstrated that the sensitivity of the 3D carbon electrodes was about three times as high as that of the 2D carbon electrodes on stripping analysis of both heavy metals. The electrochemical responses of 1 µg L-1 Pb2+ at the 3D carbon electrodes were about 6 times as high as those at the 2D carbon electrodes. The improved analytical performance of disposable 3D carbon electrodes could be attributed to their increased electrochemical effective area, which was brought by replacing SSSs with SSWM. The obtained disposable 3D carbon electrodes could be used for differentiation of Pb in teethers and corns. This study not only presented the potential of SSWM in the preparation of disposable 3D carbon electrodes but also suggested a simple and effective strategy for the preparation of disposable 3D electrodes for practical applications.

[Simultaneous determination of seven flavonoids in Nervilia fordii with HPLC].

The study is to develop an HPLC method for simultaneous determination of rhamnazin (1), rhamnocitrin (2), rhamnetin (3), rhamnazin-3-O-beta-D-glucopyranoside (4), rhamnazin-3-O-beta-D-xylopyranosyl-(1-->4)-beta-D-glucopyranoside (5), rhamnazin-3-O-beta-D-glucopyranosyl-(1-->4)-beta-D-glucopyranoside (6), and rhamnocitrin-3-O-beta-D-glucopyranosyl-(1-->4)-beta-D-glucopyranoside (7) in Nervilia fordii. The separation was performed on a Kromasil C18 column (250 mm x 4.6 mm, 5 microm) with 0.4% phosphoric acid-acetonitrile as the mobile phase in a gradient elution at a flow rate of 1.0 mL x min(-1). The detect wavelength was set at 256 nm, and the column temperature was set at 40 degrees C. There were good linear relationships between the logarithm values of concentrations and those of the peak areas of seven flavonoids (1-7) in the range of 0.55-70.00 microg x mL(-1) (r = 0.9997), 0.86-110.00 microg x mL(-1) (r = 0.9997), 0.39-50.00 microg x mL(-1) (r = 0.999 7), 0.55-70.00 microg x mL(-1) (r = 0.999 5), 1.33-170.00 microg x mL(-1) (r = 0.9998), 1.33-170.00 microg x mL(-1) (r = 0.9998), 0.16-20.00 microg x mL(-1) (r = 0.9995), respectively. The recoveries of the seven flavonoids were between 97.19%-99.45%, the relative standard deviations (RSDs) were between 0.91%-2.69%. The established method is rapid, accurate with high repeatability, which could provide scientific evidence for the quality control of Nervilia fordii.

Two new acetylated flavan glycosides from rhizomes of the fern Abacopteris penangiana.

Two new acetylated flavan glycosides (2S,4R)-4,5,7-trihydroxy-4'-methoxy-6,8-dimethylflavan-5-O-ß-D-6-acetylglucopyranoside-7-O-ß-D-glucopyranoside (1) and (2S,4R)-5,7-dihydroxy-4,4'-dimethoxy-6,8-dimethylflavan-5-O-ß-D-6-acetylglucopyranoside-7-O-ß-D-glucopyranoside (2) were isolated from the rhizomes of Abacopteris penangiana. Their structures were elucidated on the basis of spectroscopic methods including IR, HR-ESI-MS, 1D and 2D NMR, and chemical evidences.

[Analysis of the chemical constituents of essential oil from the leaves of Callicarpa integerrima].

OBJECTIVE: To analyze the compositions of the essential oil from the leaves of Callicarpa integerrima. METHODS: GC-MS method was used. RESULTS: 42 chemical constituents, accounting for 97.62% of total content were identified. CONCLUSION: The main components were beta-caryophyllene (33.74%), Elixene (12.86%), tau-Cadinene (9.57%) and (-) -Spathulenol (8.99%). Besides, some sesquiterpenes and their oxides, such as Copaene (4.21%), Globulol (3.81%), alpha-Caryophyllene (2.48%), 2-Naphthalenemethanol, decahydro-alpha, alpha, 4a-trimethyl-8-methylene-[2R-(2alpha, 4aalpha, 8alphabeta)]-(2.37%) and 1,6-Cyclodecadiene (2.24%) had also relatively high contents.

[Studies on the chemical constituents of Callicarpa kochiana].

OBJECTIVE: To study on the chemical constituents of Callicarpa kochiana. METHODS: The chemical constituents were isolated by chromatographic methods and structurally elucidated by spectral analysis. RESULTS: Twelve compounds were obtained and identified as alpha-amyrin(I), 2beta, 3beta, 19alpha-trihydroxy-12-en-28-ursolic acid (II), oleanolic aicd (III), alpha-amyrin-3-0-beta-D-glucopyranoside (IV), ursolic acid (V), betulinic acid (VI), 2alpha, 3beta,23-trihydroxy-12-en-28-oic-0-beta-D-glucopyranoside (VII), 0-hydroxybenzoic acid (VI), pomolic acid (IX), myrianthic acid (X), beta-sitosterol (XI), dauricine (XII). CONCLUSION: All of these compounds are isolated from Callicarpa kochiana for the first time and compounds II, IV, VII, VIII, IX and X are reported for the first time from Callicarpa genus.

Paper-based electroanalytical devices for stripping analysis of lead and cadmium in children's shoes.

Children's shoes are potential sources of toxic heavy metals, especially for younger children. Electrochemical detection could be applied for effective stripping analysis of heavy metals (such as Cd and Pb). However, the substrates of working electrodes are still limited and it is not well known which property is critical. Herein ITO glass was used as the substrate and the working electrode was modified with carbon cement for stripping analysis of Cd and Pb. The electrochemical impedance spectra of the ITO modified electrodes suggested the connection between the resistance and the electrochemical responses of heavy metals in stripping analysis, depending on the dilution ratio of the carbon cement. After optimization, the ITO modified electrodes in paper-based analytical devices could be used to sensitively quantify Cd and Pb with the concentration ranging from 10 to 1000 ppb. The detection limit of Pb2+ could reach less than 1 ppb while that of Cd2+ could reach 5 ppb, depending on the pH value of the sample solution. The paper-based electroanalytical devices could be used to quantify the concentration of Cd and Pb in children's shoes. This study implied the impact of the electric conductivity of the electrode substrates on stripping analysis, which might help to find more materials for the fabrication of the working electrodes.

New glycosylsphingolipids from Psychotria serpens L.

In clinical, Psychotria serpens L. was often substitute for Caulis trachelospermi to treat cancer in China. Meanwhile, EtOAc and n-BuOH fractions of MeOH extract of P. serpens L. show power activity against H460, HepG2, Hela, and PC9/GR cell lines, and no toxic effects against normal 16HBE cell lines. In our ongoing search for bioactive novel compounds from Chinese material medica, one new type of glycosylsphingolipids Psychotramide (1a-1c) were isolated from P. serpens L., and their structures were identified through spectroscopic techniques including NMR (1D and 2D) and MS (LC-MS, and GC-MS).

New amide alkaloids from Delphinium brunonianum.

Five new amide alkaloids, named delamide A-E (1-5), along with five known ones, methyl-N-(3-carboxy-2-methylpropanoyl) anthranilate (6), benzoic acid, 2-[(1-oxodecyl) amino]-methylester (7), puberline (8), benzoic acid, 2-[(4-methoxy-2-methyl-1, 4-dioxobutyl) amino]-methylester (9) and benzoic acid, 2-[(4-methoxy-3-methyl-1, 4-dioxobutyl) amino]-methylester (10) were isolated from the extract of Delphinium brunonianum. Their structures were elucidated by extensive spectroscopic analyses (including 1D-, 2D-NMR, and HR-ESI-MS). 1-10 were also evaluated for their acetylcholinesterase (AChE) inhibiting activity by the Ellman's method. Delamide A (1) showed highly selective AChE inhibition activity. The kinetic analysis revealed that 1 was a mixed-type reversible inhibitor of AChE.

Low-risk gestational trophoblastic neoplasia outcome after treatment with VMP regimen from 2005 to 2017.

OBJECTIVE: To evaluate the efficacy and toxicity of VMP regimen applied to the patients with low-risk gestational trophoblastic neoplasia (LR-GTN) treated in Anhui provincial hospital. MATERIALS AND METHODS: Between 2005 and 2017, 87 patients with low-risk gestational trophoblastic neoplasia received VMP regimen, consisted of vincristine (VCR), methotrexate (MTX) and platinum (cisplatin, carboplatin or nedaplatin), 68 of whom received VMP as their first-line chemotherapy, and 19 methotrexate-failed patients received VMP regimen as their second-line chemotherapy. The staging and scoring system was based on International Federation of Gynecology and Obstetrics (FIGO 2000) criteria. We describe and analyze their baseline characteristics, remission/resistance/recurrence rates, adverse reactions and prognosis. RESULTS: The first-line VMP protocol can achieve an 83.8% remission rate and it tended to develop resistance when the pretreatment ß-hCG reaches 7503.5 IU/L, and can achieve complete remission with FAV and EMA-CO as the salvage regimen. Among the 19 methotrexate-failed patients, 2 of whom were yet resistant to VMP regimen, followed by several courses of salvage chemotherapy such as FAV and EMP, and achieved 89.5% remission rate in second-line VMP group. Resistance to this regimen was obviously related with higher pre-treatment HCG whether used as primary or salvage treatment. Severe myelosuppression (grade 3 or 4) was shown in 4 (5.9%) of 68 cases, of which none was grade 4. CONCLUSION: For patients diagnosed with LR-GTN VMP regimen was a safe and effective treatment with a high rate of remission.

Whole exome sequencing identifies a rare variant in DAAM2 as a potential candidate in idiopathic pulmonary ossification.

BACKGROUND: Diffuse pulmonary ossification (DPO) is a rare disease characterized by bone tissue formation in the lung. DPO can be classified into idiopathic pulmonary ossification (IPO) and secondary pulmonary ossification. Cases with no identified etiology are classified as IPO. Variants of dishevelled associated activator of morphogenesis 2 (DAAM2) have been reported to be involved in the bone-resorption of osteoclasts. METHODS: Whole exome sequencing (WES) was used on samples from a patient with IPO and his healthy parents. The effects of all variants were determined using functional predictors (PolyPhen-2, SIFT, FATHMM and MutationTaster); variants existing only in the patient were further screened compared with his healthy parents. RESULTS: Forty deleterious variants, including 25 single nucleotide variants (SNVs) and 15 insertions and deletions (indels), were identified by WES. Finally, DAAM2 (c.G2960T:p.R987L) was screened by pathway analysis. CONCLUSIONS: We identified a novel variant of DAAM2 (c.G2960T:p.R987L) that might participate in the disease process of IPO.

[Two new flavan glycosides from Abacopteris penangiana].

To study the chemical constituents of Abacopteris penangiana (Hook.) Ching, various chromatographic techniques were used to isolate and purify the constituents. The structures of the obtained compounds were elucidated by spectroscopic data and physical-chemical properties. Two compounds were isolated from the n-BuOH soluble fraction of an acetone-H2O (4:1) extract of A. penangiana and were identified as 4'-hydroxy pneumatopterin B (I) and 6"-O-acetyl triphyllin A (II). Compounds I and II are new compounds.