Effect of mPEG-PLGA on Drug Crystallinity and Release of Long-Acting Injection Microspheres: In Vitro and In Vivo Perspectives.

PURPOSE: Traditional progesterone (PRG) injections require long-term administration, leading to poor patient compliance. The emergence of long-acting injectable microspheres extends the release period to several days or even months. However, these microspheres often face challenges such as burst release and incomplete drug release. This study aims to regulate drug release by altering the crystallinity of the drug during the release process from the microspheres. METHODS: This research incorporates methoxy poly(ethylene glycol)-b-poly(lactide-co-glycolide) (mPEG-PLGA) into poly(lactide-co-glycolide) (PLGA) microspheres to enhance their hydrophilicity, thus regulating the release rate and drug morphology during release. This modification aims to address the issues of burst and incomplete release in traditional PLGA microspheres. PRG was used as the model drug. PRG/mPEG-PLGA/PLGA microspheres (PmPPMs) were prepared via an emulsification-solvent evaporation method. Scanning electron microscopy (SEM), powder X-ray diffraction (PXRD), and differential scanning calorimetry (DSC) were employed to investigate the presence of PRG in PmPPMs and its physical state changes during release. RESULTS: The addition of mPEG-PLGA altered the crystallinity of the drug within the microspheres at different release stages. The crystallinity correlated positively with the amount of mPEG-PLGA incorporated; the greater the amount, the faster the drug release from the formulation. The bioavailability and muscular irritation of the long-acting injectable were assessed through pharmacokinetic and muscle irritation studies in Sprague-Dawley (SD) rats. The results indicated that PmPPMs containing mPEG-PLGA achieved low burst release and sustained release over 7 days, with minimal irritation and self-healing within this period. PmPPMs with 5% mPEG-PLGA showed a relative bioavailability (Frel) of 146.88%. IN CONCLUSION: In summary, adding an appropriate amount of mPEG to PLGA microspheres can alter the drug release process and enhance bioavailability.

Pharmacokinetics and molecular docking of the cardioprotective flavonoids in Dalbergia odorifera.

The purpose of this research was to investigate the cardioprotective effects and pharmacokinetics of Dalbergia odorifera flavonoids. The cardioprotective effects were detected by hematoxylin-eosin staining histopathological observations and the detection of myocardial enzymes by kits in serum, peroxidation and antioxidant levels and ATPase activities by kits in the homogenate supernatant, and antioxidant and apoptosis-related protein expression in heart tissue by immunohistochemistry. The pharmacokinetics parameters of the flavonoids in rat plasma were investigated by ultra-high-performance liquid chromatography coupled with tandem mass spectrometry. Molecular docking of the compounds absorbed by the blood with specific proteins was carried out. D. odorifera flavonoids significantly reduced the levels of creatinine kinase, alanine transaminase, nitric oxide, and Hydrogen peroxide, elevated the levels of glutathione, superoxide dismutase, and ATPase, significantly reduced the pathological degree of heart tissue and had obvious anti-myocardial ischemia efficacy. Nine out of the 17 flavonoids were detected in rat plasma. The peak concentration and the area under the plasma concentration-time curve values of 3'-O-methylviolanone and sativanone were significantly higher than those of other ingredients. The peak time of most flavonoids (except for Genistein and Pruneion) was lower than 2 h, while the half-life of elimination of the nine flavonoids ranged from 3.32 to 21.5 h. The molecular docking results showed that daidzein, dalbergin, formononetin, and genistein had the potential to bind to the target proteins. The results of the study provide an important basis for understanding the cardioprotective effects and clinical application of D. odorifera.

[Anxiolytic and antidepressant effects of agarwood inhalation and its mechanism].

This study used m-chloropheniperazine(MCPP) and chronic unforeseeable mild stress(CUMS) to induce the rat models of anxiety and depression, respectively. The behaviors of rats were observed by the open field test(OFT), light-dark exploration test(LDE), tail suspension test(TST), and forced swimming test(FST), and the antidepressant and anxiolytic effects of agarwood essential oil(AEO), agarwood fragrant powder(AFP), and agarwood line incense(ALI) were explored. The enzyme-linked immunosorbent assay(ELISA) was used to determine the levels of 5-hydroxytryptamine(5-HT), glutamic acid(Glu), and γ-aminobutyric acid(GABA_A) in the hippocampal area. The Western blot assay was used to determine the protein expression levels of glutamate receptor 1(GluR1) and vesicular glutamate transporter type 1(VGluT1), exploring the anxiolytic and antidepressant mechanism of agarwood inhalation. The results showed that compared with the anxiety model group, the AEO, AFP, and ALI groups decreased the total distance(P<0.05), decreased the velocity of movements(P<0.05), prolonged the immobile time(P<0.05), and reduced the distance and velocity of the rat model of anxiety in the dark box(P<0.05). Compared with the depression model group, the AEO, AFP, and ALI groups increased the total distance and average velocity(P<0.05), reduced the immobile time(P<0.05), and reduced the forced swimming and tail suspension time(P<0.05). In terms of transmitter regulation, the AEO, AFP, and ALI groups decreased the level of Glu in the rat model of anxiety(P<0.05) and increased the levels of GABA_A and 5-HT(P<0.05), while the AEO, AFP, and ALI groups all increased the level of 5-HT in the rat model of depression(P<0.05) and decreased the levels of GABA_A and Glu(P<0.05). At the same time, the AEO, AFP, and ALI groups all increased the protein expression levels of GluR1 and VGluT1 in the hippocampus of the rat models of anxiety and depression(P<0.05). In conclusion, AEO, AFP, and ALI exert anxiolytic and antidepressant effects, and the mechanism might be related to the regulation of the neurotransmitter and the protein expression of GluR1 and VGluT1 in the hippocampus.

Characterization of femtosecond laser-induced grating scattering of a continuous-wave laser light in air.

Nanosecond laser-induced grating scattering/spectroscopy (LIGS) technique has been widely applied for measuring thermodynamic parameters such as temperature and pressure in gaseous and liquid media. Recently, femtosecond (fs) laser was demonstrated to induce the grating and develop the fs-LIGS technique for gas thermometry. In this work, we systematically investigated the fs-LIGS signal generation using 35 fs, 800 nm laser pulses at 1 kHz repetition rate in ambient air by varying the pump laser energies, the probe laser powers and the temporal delays between two pump laser pulses. The stability of single-shot fs-LIGS signal was studied, from which we observed that the signal intensity exhibits a significant fluctuation while the oscillation frequency shows a much better stability. A 4.5% precision of the oscillation frequency was achieved over 100 single-shot signals. By using a previously-developed empirical model, the fs-LIGS signals were fitted using nonlinear least-squares fitting method, by which crucial time constants characterizing the signal decay process were extracted and their dependences on the pump laser energy were studied. From the measured results and theoretical analysis, we found that the appropriate range of the overall pump laser energy for reliable fs-LIGS measurements is approximately located within 80 ∼ 300 µJ. The limitations on the accuracy and precision of the fs-LIGS measurements, the origin of destructive influence of plasma generation on the signal generation as well as the electrostriction contribution were also discussed. Our investigations could contribute to a better understanding of the fs-LIGS process and further applications of the technique in single-shot gas thermometry and pressure measurements in various harsh conditions.

Gas-phase pressure measurement using femtosecond laser-induced grating scattering technique.

Gas-phase pressure measurements remain challenging in situations where local pressure rapidly changes or in hostile environments such as turbulent combustion. In this work, we demonstrate the implementation of the recently developed femtosecond laser-induced grating scattering (fs-LIGS) technique for pressure measurement in ambient air. With an overall femtosecond laser pulse energy of 185 µJ, fs-LIGS signals were generated for various gas pressure ranging from 0.2 to 3.0 bar. By theoretically fitting the signal and extracting the time constant of the stationary density modulation damping, the pressure is successfully derived. The derived values were compared to the gauge pressure, which shows a quasi-linear dependence with a slope of 0.96, suggesting the feasibility of the fs-LIGS technique for gas-phase pressure measurements.

Effective Components and Molecular Mechanism of Agarwood Essential Oil Inhalation and the Sedative and Hypnotic Effects Based on GC-MS-Qtof and Molecular Docking.

Agarwood has been used for the administration of hypnotic therapy. Its aromatic scent induces a relaxed state. However, its aromatic constituents and the underlying molecular effect are still unclear. This study aims to determine the active substance and molecular mechanism of the hypnotic effect of agarwood essential oil (AEO) incense inhalation in insomniac mice. Insomnia models were induced by para-chlorophenylalanine (PCPA, 300 mg/kg) in mice. The sleep-promoting effect was evaluated. Neurotransmitter levels and its receptor were detected to explore the molecular mechanism. The effective components were analyzed by GC-Q/TOF-MS of AEO. The binding mechanisms of the core compounds and core targets were verified by molecular docking. These results showed that AEO inhalation could significantly shorten sleep latency and prolong sleep time, inhibit autonomous activity and exert good sedative and sleep-promoting effects. A mechanistic study showed that AEO inhalation increased the levels of γ-aminobutyric acid (GABAA), the GABAA/glutamic acid (Glu) ratio, 5-hydroxytryptamine (5-HT) and adenosine (AD), upregulated the expression levels of GluR1, VGluT1 and 5-HT1A and downregulated 5-HT2A levels. Component analysis showed that the most abundant medicinal compounds were eremophilanes, cadinanes and eudesmanes. Moreover, the docking results showed that the core components stably bind to various receptors. The study demonstrated the bioactive constituents and mechanisms of AEO in its sedative and hypnotic effects and its multicomponent, multitarget and multipathway treatment characteristics in PCPA-induced insomniac mice. These results provide theoretical evidence for insomnia treatment and pharmaceutical product development with AEO.

[Anti-asthmatic effect of agarwood alcohol extract in mice and its mechanism].

As recorded, agarwood has the function of improving qi reception and relieving asthma, but the underlying mechanism is unclear and rarely reported. Therefore, this study explored the anti-asthmatic effect of the alcohol extract of agarwood produced by the whole-tree agarwood-inducing technique(Agar-Wit) in the asthma mouse model induced by intraperitoneal injection of ovalbumin(OVA) + Al(OH)_3 combined with intranasal administration of OVA and the mechanism, and compared the anti-asthmatic effects of agarwood induced with different methods. Firstly, the anti-inflammatory and anti-asthmatic effects of Agar-Wit agarwood in mice were evaluated based on the asthma frequency, lung tissue injury, and peripheral inflammatory white blood cell(WBC) count and eosinophil count. Then, the levels of interleukin-1ß(IL-1ß), IL-17, and IL-10 in serum of mice were detected by enzyme-linked immunoassay(ELISA) and the expression of inflammation-and apoptosis-related genes in tissues was measured by reverse transcription polyme-rase chain reaction(RT-PCR) so as to preliminarily explore the anti-asthmatic mechanism. RESULTS:: showed that the alcohol extract of Agar-Wit agarwood significantly reduced asthma frequency, relieved pathological injury, improved peripheral WBC count and eosinophil count, decreased the levels of inflammatory cytokines IL-1ß and IL-17, elevated the level of anti-inflammatory cytokine IL-10, and down-regulated the mRNA expression of IL-1 R, tumor necrosis factor receptor R(TNFR), nuclear transcription factor-kappa B(NF-κB), Bax, and caspase 3, but had no significant influence on the expression of high-mobility group box 1(HMGB1) protein, caspase 8, and Bcl-2. The effect of Agar-Wit agarwood alcohol extract was better than that of wild agarwood alcohol extract and alcohol extract of agarwood induced with the burning-chisel-drilling method at the same dose. In conclusion, Agar-Wit agarwood can significantly alleviate inflammation and asthma, which is related to its anti-inflammation and anti-apoptosis activity.

In Vivo Structural and Functional Abnormalities of the Striatums Is Related to Decreased Astrocytic BDNF in Itpr2-/- Mice Exhibiting Depressive-Like Behavior.

Background: Previous researches indicate that Itpr2 -/- mice (inositol 1,4,5-trisphosphate receptor type 2 knockout mice) show depressive-like symptoms; however, little is known regarding the in vivo neurobiological effect of Itpr2 as well as the specific pattern of brain abnormalities in Itpr2 -/- mice. Methods/Materials. First, behavioral tests, structural magnetic resonance imaging (MRI), and resting-state functional MRI were performed on Itpr2 -/- mice and matched healthy controls. Voxel-based morphometry and seed-based voxel-wise functional connectivity (FC) were, respectively, calculated to assess the gray matter volume and the functional activities of the brain in vivo. Second, the sample of relevant changed brain regions was extracted to detect the expression of BDNF. Finally, to further validate the relationship between Itpr2 deficiency and the observed brain abnormalities, we performed Western blotting to detect the expression of pro-BDNF and mBDNF in Itpr2 -/- C8-D1A (a type of astrocyte). Results: Compared with controls, Itpr2 -/- mice showed depressive-like behaviors as well as significantly lower gray matter volume in striatums mainly, periaqueductal GM, and the right frontoparietal cortices as well as lower striatal-hippocampal and striatal-right parietal cortex (mainly for the primary and secondary somatosensory cortex) FC. Moreover, decreased expression of mBDNF was found in both sample tissues of the striatum in Itpr2 -/- mice and Itpr2 -/- C8-D1A. Conclusion: By combining biochemistry and MR analyses, this study provides evidences to support that the Itpr2-related neuropathological effect is possibly mediated by the striatal abnormality associated with dysfunctional astrocytes in Itpr2 -/- mice in vivo, thus may help us better understand underlying mechanisms of Itpr2 deficiency as well as its relation to depressive-like behavior.

Crystal structure of PppA from Pseudomonas aeruginosa, a key regulatory component of type VI secretion systems.

The Type VI secretion system (T6SS) is a membrane protein complex related to inter-bacterial competitions and host-pathogen interactions in Pseudomonas aeruginosa. The T6SS is regulated by a great variety of regulatory mechanisms at multiple levels, including post-translational modification with threonine phosphorylation mediated by Ser/Thr protein kinase PpkA and phosphatase PppA. The T6SS is activated by PpkA via Thr phosphorylation of Fha, and PppA can antagonize PpkA. PppA is a PP2C-family protein phosphatase and plays a key role in the disassembly and reassembly of T6SS organelles. Herein, we report the first crystal structure of PppA from Pseudomonas aeruginosa, which was determined at a resolution of 2.10 Å. The overall structure consists of a bacteria PPM structural core and a flexible flap subdomain. PppA harbors a catalytic pocket containing two manganese ions which correspond to the canonical dinuclear metal center of Ser/Thr protein phosphatases including the bacterial PPM phosphatases and human PP2C. The flexibility and the diversity of the sequence of flap subdomain across the homologues might provide clues for substrates specific recognition of phosphatases.

Two New Chromone Glycosides from the Roots of Saposhnikovia divaricata.

Five chromone glycosides were isolated from the water-soluble portions of 70% EtOH extract of the roots of Saposhnikovia divaricata, including two new chromone glycosides 1 and 2. The structures of the chromone glycosides were identified as (3'S)-3'-O-ß-d-apiofuranosyl-(1 → 6)-ß-d-glucopyranosylhamaudol (1), (2'S)-4'-O-ß-d-apiofuranosyl-(1 → 6)-ß-d-glucopyranosylvisamminol (2), 3'-O-glucopyranosylhamaudol (3), 4'-O-ß-d-glucopyranosylvisamminol (4), and 4'-O-ß-d-glucopyranosyl-5-O-methylvisamminol (5) on the basis of extensive spectroscopic methods, and the absolute configurations of the new compounds were elucidated by the electronic circular dichroism (ECD) calculation and acid hydrolysis. The cytotoxic activities of the glycosides 1 - 5 against three human cancer cell lines (PC-3, SK-OV-3, and H460) were evaluated. The result showed that compounds 1 - 5 had weak cytotoxic activities against the human cancer cell lines with IC50 values in the range of 48.54 ± 0.80 - 94.25 ± 1.45 µm.

The draft genome, transcriptome, and microbiome of Dermatophagoides farinae reveal a broad spectrum of dust mite allergens.

BACKGROUND: A sequenced house dust mite (HDM) genome would advance our understanding of HDM allergens, a common cause of human allergies. OBJECTIVE: We sought to produce an annotated Dermatophagoides farinae draft genome and develop a combined genomic-transcriptomic-proteomic approach for elucidation of HDM allergens. METHODS: A D farinae draft genome and transcriptome were assembled with high-throughput sequencing, accommodating microbiome sequences. The allergen gene structures were validated by means of Sanger sequencing. The mite's microbiome composition was determined, and the predominant genus was validated immunohistochemically. The allergenicity of a ubiquinol-cytochrome c reductase binding protein homologue was evaluated with immunoblotting, immunosorbent assays, and skin prick tests. RESULTS: The full gene structures of 20 canonical allergens and 7 noncanonical allergen homologues were produced. A novel major allergen, ubiquinol-cytochrome c reductase binding protein-like protein, was found and designated Der f 24. All 40 sera samples from patients with mite allergy had IgE antibodies against rDer f 24. Of 10 patients tested, 5 had positive skin reactions. The predominant bacterial genus among 100 identified species was Enterobacter (63.4%). An intron was found in the 13.8-kDa D farinae bacteriolytic enzyme gene, indicating that it is of HDM origin. The Kyoto Encyclopedia of Genes and Genomes pathway analysis revealed a phototransduction pathway in D farinae, as well as thiamine and amino acid synthesis pathways, which is suggestive of an endosymbiotic relationship between D farinae and its microbiome. CONCLUSION: An HDM genome draft produced from genomic, transcriptomic, and proteomic experiments revealed allergen genes and a diverse endosymbiotic microbiome, providing a tool for further identification and characterization of HDM allergens and development of diagnostics and immunotherapeutic vaccines.

[Macroscopic and Microscopic Identification of Ganoderma and The Confusable Medicinal Mushrooms of "Zhi"].

OBJECTIVE: To compare the macroscopic and microscopic characteristics of Ganoderma and the confusable medicinal mushrooms of "Zhi". METHODS: The methods of macroscopic and microscopic examination were employed to authenticate Ganoderma and the other medicinal mushrooms of "Zhi" in the fungal group. RESULTS: The medicinal mushroom named "Zhi" could be distinguished from each other by the shape of pileus, stipe and the characteristics of mycelium and spore. CONCLUSION: There are abundant resources of fungal group from Ganodermataceae in China. The medicinal mushrooms of "Zhi", which are quite complicated in the market, are needed to be differentiated.

[Rats hyperuricemia model established by lipid emulsion simulating irregular of diet].

Due to the irregular of diet and overfeeding greasy and surfeit flavor closely associated with hyperuricemia disease, the lipid emulsion containing high cholesterol was used to model. To obtain a more stable and sustained animal model for the efficacy evaluation of traditional Chinese herbs, we observed the influence on the serum uric acid of rat induced by the lipid emulsion compared with high purine diet. 36 SD male rats were randomized to the normal control group, high purine diet group and lipid emulsion group respectively. The general behavior, body weight and daily food intake of rats were observed. The orbital blood was taken to separate into the serum and 24 hours urine was collected. The serum indexes such as UA, BUN, Cr, ALT, AST, TC, TG, LDL-c were determined every 2 weeks, and XOD, ADA enzyme activity were determined at the 4th week. The urine indexes such as UA, Cr and Cua/Ccr were determined at the 4th week. After stopping modeling, the serum UA were determined two weeks and four weeks later respectively. At the 2nd week, the body weight and daily food intake of rats in the lipid emulsion group reduced significantly, and the level of serum UA, BUN, Cr, TC, LDL-c, ATL, AST raised significantly meanwhile TG reduced. At the 4th week, the serum UA in high purine diet group did not raise, and the serum XOD raised obviously while ADA did not; the serum UA in lipid emulsion group was higher significantly, and the serum XOD and ADA raised while Cua/Ccr reduced obviously. At the 6th weeks, the serum UA in both the high purine diet group and lipid emulsion group raised obviously. After stopping modeling, the serum UA in lipid emulsion group still maintained a high level at the 2nd week and back to the normal level at the 4th week. Compared with high purine diet, the hyperuricemia model induced by lipid emulsion forms earlierand more stable. It maybe has great value to study the pharmacodynamics of traditional Chinese medicine treatment to hyperuricemia disease. Its mechanism may be related to increasing XOD and ADA enzyme activity which can promote uric acid synthesis, meanwhile inhibiting of uric acid excretion.

[Chemical constituents of Juncus setchuensis].

OBJECTIVE: To study the chemical constituents of Juncus setchuensis. METHODS: Column chromatography was used in the isolation procedure. The structures of isolated compounds were elucidated by spectral data. RESULTS: Eight compounds were isolated and their structures were identified as 2-hydroxy-3-methylanthraquinone (1), physcion (2), stigmasterol (3), stigmast-3,6-dione (4), vanillin (5), n-heptacosanoic acid (6), trans-hydroxycinnamic (7) and 4-hydroxy-3-methoxy benzoic acid (8). CONCLUSION: Compound 1, 2, 4 and 6 are obtained from this genus for the first time and all the compounds are obtained from this plant for the first time.

Severe hyponatremia and diabetes insipidus caused by low-dose cyclophosphamide in breast cancer patients: A case report and literature review.

RATIONALE: Cyclophosphamide (CTX) is widely used in the treatment of malignancies and autoimmune diseases. Although severe hyponatremia caused by low-dose CTX chemotherapy is uncommon, it can lead to serious complications and even death. PATIENT CONCERNS: A 44-year-old woman with left-sided breast cancer suddenly experienced headaches, disorientation and weakness after receiving low-dose neoadjuvant chemotherapy combined with CTX and doxorubicin. DIAGNOSES: The patient pathology showed invasive breast carcinoma. She developed severe hyponatremia and a generalized seizure after completing the first cycle of neoadjuvant chemotherapy with CTX and doxorubicin. Laboratory tests showed a serum sodium of 118 mmol/L (normal range 135-145 mmol/L) and potassium sodium 3.16 mmol/L (normal range 3.5-5.5 mmol/L). Subsequently, the patient developed secondary diabetes insipidus 4 hours after sodium supplementation, her 24-hour urine volume was 4730 mL (normal range 1000-2000 mL/24 hours), and the urine specific gravity decreased to 1.005. INTERVENTIONS: The patient was given intravenous sodium chloride (500 mL of 3%NaCl, 100 mL/hour) and potassium chloride (500 mL of 0.3%KCl, 250 mL/hour). Meanwhile, she was advised to reduce her water intake, and pituitrin was administered to prevent dehydration caused by diabetes insipidus. OUTCOMES: The patient completely recovered after correcting of the serum sodium concentration (137 mmol/L) without any neurological deficits. After discontinuing pituitrin, her 24-hour urine volume was 2060 mL and the urine specific gravity was 1.015. LESSONS: This is a typical case of severe hyponatremia induced by low-dose CTX. Clinicians and healthcare providers should be aware of this potential toxicity, and appropriate monitoring should be implemented.

Triazoles in Medicinal Chemistry: Physicochemical Properties, Bioisosterism, and Application.

Triazole demonstrates distinctive physicochemical properties, characterized by weak basicity, various dipole moments, and significant dual hydrogen bond acceptor and donor capabilities. These features are poised to play a pivotal role in drug-target interactions. The inherent polarity of triazole contributes to its lower logP, suggesting the potential improvement in water solubility. The metabolic stability of triazole adds additional value to drug discovery. Moreover, the metal-binding capacity of the nitrogen atom lone pair electrons of triazole has broad applications in the development of metal chelators and antifungal agents. This Perspective aims to underscore the unique physicochemical attributes of triazole and its application. A comparative analysis involving triazole isomers and other heterocycles provides guiding insights for the subsequent design of triazoles, with the hope of offering valuable considerations for designing other heterocycles in medicinal chemistry.

A through-space charge transfer pyrene-based fluorophore with anti-quenching behavior for deep-blue organic light-emitting devices.

A through-space charge transfer pyrene-based fluorophore has been developed for organic light-emitting devices (OLEDs). This material exhibits deep-blue fluorescence, bipolar characteristics, and anti-quenching behavior in the solid state. It proves to be an effective emitter for both doped and nondoped deep-blue OLEDs.

Current therapy in amyotrophic lateral sclerosis (ALS): A review on past and future therapeutic strategies.

Amyotrophic lateral sclerosis (ALS) is a fatal neurodegenerative disease that affects the first and second motoneurons (MNs), associated with muscle weakness, paralysis and finally death. The exact etiology of the disease still remains unclear. Currently, efforts to develop novel ALS treatments which target specific pathomechanisms are being studied. The mechanisms of ALS pathogenesis involve multiple factors, such as protein aggregation, glutamate excitotoxicity, oxidative stress, mitochondrial dysfunction, apoptosis, inflammation etc. Unfortunately, to date, there are only two FDA-approved drugs for ALS, riluzole and edavarone, without curative treatment for ALS. Herein, we give an overview of the many pathways and review the recent discovery and preclinical characterization of neuroprotective compounds. Meanwhile, drug combination and other therapeutic approaches are also reviewed. In the last part, we analyze the reasons of clinical failure and propose perspective on the treatment of ALS in the future.

[Cloning, expression and identification of Der f7 gene from Dermatophagoides farinae and its immunological characteristics].

OBJECTIVE: To clone and express Der f7 gene of Dermatophagoides farinae, and identify its immunogenicity. METHODS: Total RNA was extracted from D. farinae mites. A reference sequence (Accession No. AY283292) was used to design specific primers. The Der f7 gene fragment was amplified by RT-PCR, and cloned into pET-32a vector. The recombinant plasmid was transformed into E. coli BL21 (DE3) and induced with IPTG for protein expression. The recombinant protein was purified by Ni2+ chelating affinity chromatography and analyzed by SDS-PAGE and Western blotting. RESULTS: The Der f7 gene fragment was about 650 bp, and shared 99% homology with the published one (Accession No. FJ436108). SDS-PAGE result showed its relative molecular weight (M(r)) of 23 000. The recombinant protein showed appropriate combination ability with IgE in sera of mite allergic patients. CONCLUSION: Der f 7 gene has been expressed in prokaryotic expression system and shows allergenicity.

Agarwood extract mitigates alcoholic fatty liver in C57 mice via anti­oxidation and anti­inflammation.

Alcoholic fatty liver disease (AFLD) is a disease with a high incidence rate among individuals who drink alcohol. Our previous study found that agarwood alcohol extracts (AAEs) have a protective effect against drug­induced liver damage via anti­inflammatory and antioxidant mechanisms. Therefore, we hypothesized that agarwood may have a protective effect against AFLD. The present study assessed the potential protective effects and the underlying mechanism of action of AAEs for the treatment of an AFL in vivo model. The AFLD mouse model was established by continuous high fat diet and alcohol gavage in C57 mice. After treatment with AAEs, blood was collected, liver and adipose tissues were removed and liver and adipose indexes were analyzed. The levels of alanine aminotransferase (ALT), aspartate aminotransferase (AST), triglyceride (TG) and cholesterol (CHO) in serum were detected. The liver tissue was assessed using pathological sections. Biochemical methods were used to detect the levels of oxidative stress in the supernatant of liver tissue homogenate. The levels of pro­inflammatory cytokines in the serum were detected by ELISA. The protein expression levels of nuclear erythroid 2­related factor 2 (Nrf2) and nuclear factor kappa­B (NF­κB) in liver tissues were detected using western blotting. AAE treatment decreased the liver and adipose indexes, reduced the levels of AST, ALT, TG and CHO, improved the liver pathological characteristics and enhanced antioxidant and anti­inflammatory activities. In addition, AAEs increased the protein expression level of Nrf2 and decreased the protein expression level of NF­κB compared with AFL mice. AAE­treated animals exhibited reduced metabolic enzyme and blood lipid levels, demonstrated improved liver function and relieved the pathological damage of AFLD induced by consuming a high fat and alcohol diet. AAEs have potential protective effects in AFLD via antioxidant and anti­inflammatory mechanisms.