Potential low toxic alternative for Na-Cl cotransporter inhibition: A diuretic effect and mechanism study of Pyrrosia petiolosa.

BACKGROUND: Hydrochlorothiazide, a diuretic commonly used for the treatment of hypertension, is often associated with serious metabolic side effects. Pyrrosia petiolosa (Christ) Ching is a traditional Chinese medicine that possesses diuretic properties, without any obvious side effects. AIM: To evaluate the diuretic effect of P. petiolosa (Christ) Ching and to elucidate its underlying mechanism of action. METHODS: Extracts obtained from different polar components of P. petiolosa (Christ) Ching were analyzed for toxicity in a Kunming mouse model. The diuretic effects of the extracts were compared to that of hydrochlorothiazide in rats. In addition, compound isolation procedures, cell assays of Na-Cl cotransporter inhibition and rat diuretic test of monomeric compounds were conducted to identify the active ingredients in the extract. Subsequently, homology modeling and molecular docking were performed to explain the reason behind the diuretic activity observed. Finally, LC-MS analysis was used to elucidate the underlying mechanism of action of P. petiolosa (Christ) Ching. RESULTS: No toxicity was observed in mice administered P. petiolosa (Christ) Ching extracts. The ethyl acetate fraction showed the most significant diuretic effect. Similar results were obtained during the analysis for Na+ content in rat urine. Further separation of P. petiolosa (Christ) Ching components led to the isolation of methyl chlorogenate, 2',3'-dihydroxy propyl pentadecanoate, and ß-carotene. Results from cell assays showed that the Na-Cl cotransporter inhibitory activity of methyl chlorogenate was greater than that of hydrochlorothiazide. This result was again confirmed by the diuresis tests of monomeric compounds in rats. The molecular simulations explain the stronger interactions between the methyl chlorogenate and Na-Cl cotransporter. Of the compounds determined using LC-MS analysis, 185 were identified to be mostly organic acids. CONCLUSIONS: P. petiolosa possesses significant diuretic activities without any obvious toxicity, with least two possible mechanisms of action. Further study on this herb is warranted.

Characterization of ovarian tissue oocytes from transgender men reveals poor calcium release and embryo development, which might be overcome by spindle transfer.

STUDY QUESTION: Can spindle transfer (ST) overcome inferior embryonic development of in vitro matured ovarian tissue oocytes (OTO-IVM) originating from testosterone-treated transgender men? SUMMARY ANSWER: ST shows some potential to overcome the embryo developmental arrest observed in OTO-IVM oocytes from transgender men. WHAT IS KNOWN ALREADY: OTO-IVM is being applied as a complementary approach to increase the number of oocytes/embryos available for fertility preservation during ovarian tissue cryopreservation in cancer patients. OTO-IVM has also been proposed for transgender men, although the potential of their oocytes remains poorly investigated. Currently, only one study has examined the ability of OTO-IVM oocytes originating from transgender men to support embryo development, and that study has shown that they exhibit poor potential. STUDY DESIGN, SIZE, DURATION: Both ovaries from 18 transgender men undergoing oophorectomy were collected for the purposes of this study, from November 2020 to September 2022. The patients did not wish to cryopreserve their tissue for fertility preservation and donated their ovaries for research. All patients were having testosterone treatment at the time of oophorectomy and some of them were also having menses inhibition treatment. PARTICIPANTS/MATERIALS, SETTING, METHODS: Sibling ovaries were collected in either cold or warm medium, to identify the most optimal collection temperature. Cumulus oocyte complexes (COCs) from each condition were isolated from the ovarian tissue and matured in vitro for 48 h. The quality of OTO-IVM oocytes was assessed by calcium pattern releasing ability, embryo developmental competence following ICSI, and staining for mitochondrial membrane potential. In vitro matured metaphase I (MI) oocytes, germinal vesicle (GV) oocytes, and in vivo matured oocytes with aggregates of smooth endoplasmic reticulum (SERa) were donated from ovarian stimulated women undergoing infertility treatment and these served as Control oocytes for the study groups. ST was applied to overcome poor oocyte quality. Specifically, enucleated mature Control oocytes served as cytoplasmic recipients of the OTO-IVM spindles from the transgender men. Embryos derived from the different groups were scored and analysed by shallow whole genome sequencing for copy number variations (CNVs). MAIN RESULTS AND THE ROLE OF CHANCE: In total, 331 COCs were collected in the cold condition (OTO-Cold) and 282 were collected in the warm condition (OTO-Warm) from transgender men. The maturation rate was close to 54% for OTO-Cold and 57% for OTO-Warm oocytes. Control oocytes showed a calcium releasing ability of 2.30 AU (n = 39), significantly higher than OTO-Cold (1.47 AU, P = 0.046) oocytes (n = 33) and OTO-Warm (1.03 AU, P = 0.036) oocytes (n = 31); both values of calcium release were similar between the two collection temperatures. Mitochondrial membrane potential did not reveal major differences between Control, OTO-Warm, and OTO-Cold oocytes (P = 0.417). Following ICSI, 59/70 (84.2%) of Control oocytes were fertilized, which was significantly higher compared to 19/47 (40.4%) of OTO-Cold (P < 0.01) and 24/48 (50%) of OTO-Warm oocytes (P < 0.01). In total, 15/59 (25.4%) blastocysts were formed on Day 5 in the Control group, significantly higher than 0/19 (0%) from the OTO-Cold (P = 0.014) and 1/24 (4.1%) in OTO-Warm oocytes (P = 0.026). Application of ST rescued the poor embryo development, by increasing the Day 5 blastocyst rate from 0% (0/19) to 20.6% (6/29) (P = 0.034), similar to that in the ICSI-Control group (25.4%, 15/59). A normal genetic profile was observed in 72.7% (8/11) of OTO-Cold, 72.7% (8/11) of OTO-Warm and 64.7% (11/17) of Control Day 3-Day 5 embryos. After ST was applied for OTO-IVM oocytes, 41.1% (7/17) of the embryos displayed normal genetic patterns, compared to 57.1% (4/7) among ST-Control Day 3-Day 5 embryos. LARGE SCALE DATA: N/A. LIMITATIONS, REASONS FOR CAUTION: Due to the limited access to human oocytes and ovarian tissue, our results should be interpreted with some caution, as only a limited number of human oocytes and embryos could be investigated. WIDER IMPLICATIONS OF THE FINDINGS: The results of this study, clearly indicate that OTO-IVM oocytes originating from transgender patients are of inferior quality, which questions their use for fertility preservation. The poor quality is likely to be related to cytoplasmic factors, supported by the increased blastocyst numbers following application of ST. Future research on OTO-IVM from transgender men should focus on the cytoplasmic content of oocytes or supplementation of media with factors that promote cytoplasmic maturation. A more detailed study on the effect of the length of testosterone treatment is also currently missing for more concrete guidelines and guidance on the fertility options of transgender men. Furthermore, our study suggests a potentially beneficial role of experimental ST in overcoming poor embryo development related to cytoplasmic quality. STUDY FUNDING/COMPETING INTEREST(S): A.C. is a holder of FWO grants (1S80220N and 1S80222N). A.B. is a holder of an FWO grant (1298722N). B.H. and A.V.S. have been awarded with a special BOF (Bijzonder Onderzoeksfonds), GOA (Geconcerteerde onderzoeksacties) and 2018000504 (GOA030-18 BOF) funding. B.H. has additional grants from FWO-Vlaanderen (Flemish Fund for Scientific Research, G051516N and G1507816N) and Ghent University Special Research Fund (Bijzonder Onderzoeksfonds, BOF funding (BOF/STA/202109/005)), and has been receiving unrestricted educational funding from Ferring Pharmaceuticals (Aalst, Belgium). The authors declare that they have no conflict of interest. TRIAL REGISTRATION NUMBER: N/A.

Suitability of botanical extracts as components of complex mixtures used in herbal tea infusions-challenges and opportunities.

Herbal tea is a mainstay dosage form in practically all systems of traditional medicine and widely used in modern alternative and complementary medicine. Incorporating botanical extracts into herbal tea formulations is of vital interest to manufacturers as it allows for the use of herbal ingredients that would otherwise not be suitable for the dosage form, for instance, dosing requirements, solubility in water, sensory constraints etc. Furthermore, reducing the amount of ingredients in a formula increases compliance with dosing recommendations and thus therapeutic benefit. However, formulating with botanical extracts comes with challenges, ranging from sourcing ingredients of appropriate quality, developing suitable methods for quality control with combinations of (herbal) ingredients, processing constraints such as hygroscopicity, solubility, dispersibility, homogeneity of distribution, and packaging machinability, all the way to stability required for hot-water infusion. We report on experiences with overcoming such challenges in a set of examples and provide guidance to the extract industry on how to tap into the bagged tea sector with better suited or tailor-made solutions for the formulator.

[IgG4-related diseases of retroperitoneum in urinary and male reproductive system: a clinicopathological analysis of eleven cases].

Objective: To analyze the clinicopathological features of IgG4-related diseases (RD) of retroperitoneum and the urinary and male reproductive system (IgG4-RUMR). Methods: A total of 11 IgG4-RUMR cases from January 2013 to March 2021 were retrospectively collected at Peking University Third Hospital and Shandong Provincial Hospital affiliated to Shandong First Medical University. The clinicopathologic features, laboratory and imaging findings were analyzed and scored according to the 2019 ACR/EULAR classification criteria for IgG4-RD. Results: The 11 patients (male:female is 9∶2; mean age 59 years, range from 44 to 83 years) were initially admitted to the Deparment of Urology/Kidney Transplantation (10 cases) and the Department of Oncology (1 case). All patients had urogenital disorders or imaging abnormalities. Three of the 11 patients had a history of IgG4-RD such as lacrimal gland engorgement, salivary gland engorgement and IgG4-associated pancreatitis. Abnormal retroperitoneal soft tissue and hydronephrosis were found in eight cases, while epididymal and spermatic cord masses were found in one case, simple renal mass in one case, and"benign prostatic hyperplasia"in one case. In the 10 patients tested for serum IgG4, the serum IgG4 level was 0.8-14.4 g/L. Histologically, all cases showed significant lymphoplasmacytic infiltration and storiform fibrosis, and some were accompanied by obliterative phlebitis. The number of IgG4 positive plasma cells was 12-155 per high-power field, and the IgG4/IgG ratio was 15%-77%. According to the 2019 ACR/EULAR IgG4-RD classification standard 11 cases scored 20-48 points, all of which met the diagnostic criteria of IgG4-RUMR. Therapeutically, the patient with a simple renal mass underwent partial nephrectomy. The patient with prostate lesion underwent transurethral resection of prostate and was initially diagnosed as nonspecific chronic prostatitis. Later, the patient was admitted again because of salivary gland swelling, and the pathologic diagnosis was amended. The patient with epididymal and spermatic cord masses participated in a clinical trial about retroperitoneal fibrosis. The remaining eight patients received symptomatic treatment such as adhesiolysis and stent placement. All the patients were subsequently treated with glucocorticoid/immunosuppressant and symptoms relieved. Conclusions: IgG4-RUMR is uncommon. In clinical practice, information from clinical, serologic, radiologic and pathologic evaluations must be integrated. IgG4-RUMR should be considered in the differential diagnosis of urinary and male reproductive diseases. The 2019 ACR/EULAR classification criteria for IgG4-RD, while relatively complex, are objective and practical in the diagnosis of IgG4-RUMR.

[A multicenter randomized prospective study of concurrent chemoradiation with 60 Gy versus 50 Gy for inoperable esophageal squamous cell carcinoma].

Objective: To determine whether 60 Gy is superior to standard 50 Gy for definitive concurrent chemoradiation(CCRT) in esophageal squamous cell carcinoma (ESCC) using modern radiation technology in a phase Ⅲ prospective randomized trial. Methods: From April 2013 to May 2017, 331 patients from 22 hospitals who were pathologically confirmed with stage ⅢA-ⅣA ESCC were randomized to 60 Gy or 50 Gy with random number table. Total of 305 patients were analyzed, including 152 in 60 Gy group and 153 in 50 Gy group. The median age was 63 years, 242(79.3%) males and 63(20.7%) females. The median length of primary tumor was 5.6 cm. The clinical characteristics between two groups were comparable. All patients were delivered 2 Gy per fraction, 5 fractions per week. Concurrent weekly chemotherapy with docetaxel (25 mg/m(2)) and cisplatin (25 mg/m(2)) and 2 cycles consolidation chemotherapy with docetaxel (70 mg/m(2)) and cisplatin (25 mg/m(2), d1-3) were administrated. The primary endpoint was local/regional progression-free survival (LRPFS). The data were compared with Pearson chi-square test or Fisher's exact test. Results: At a median follow-up of 27.3 months, the disease progression rate was 37.5% (57/152), 43.8% (67/153) in the high and standard-dose group, respectively (χ(2)=1.251, P=0.263). The 1, 2, 3-year LRPFS rate was 75.4%, 56.8%, 52.1% and 74.2%, 58.4%, 50.1%, respectively (HR: 0.95, 95%CI: 0.69-1.31, P=0.761). The 1, 2, 3-year overall survival rate was 84.1%, 64.8%, 54.1% and 85.4%, 62.9%, 54.0%, respectively (HR: 0.98, 95%CI: 0.71-1.38, P=0.927). The 1, 2, 3-year progression-free survival rate was 70.8%, 54.2%, 48.5% and 65.5%, 51.9%, 45.1%, respectively (HR: 0.93, 95%CI: 0.68-1.26, P=0.621). The incidence rates in toxicities between the two groups were similar except for higher rate of severe pneumonitis in high dose group (χ(2)=11.596, P=0.021). Conclusions: The efficacy in disease control is similar between 60 Gy and 50 Gy using modern radiation technology concurrent with chemotherapy for ESCC. The 50 Gy should be recommended as the regular radiation dose with CCRT for ESCC.

Doubly guaranteed mechanism for pollination and fertilization in Ipomoea purpurea.

Flowers, the reproductive organs of angiosperms, show a high degree of diversity in morphological structure and flowering habit to ensure pollination and fertilization of the plants. Effect of flower movement on pollination and fertilization was investigated in Ipomoea purpurea (Convolvulaceae) in this study. Fluorescence microscopy was used to observe the germination of pollen grains at different temperatures. From 04:00 to 06:00 h, the stigma was taller than the filaments, so that self-pollination could not occur, and cross-pollination was carried out by insects. Pollen grains germinated rapidly after falling on the stigma; the pollen tube reached the ovule to complete fertilization after 2-3 h. From 07:00 to 09:00 h, filaments of two stamens grew rapidly and reached the same height as the stigma, thus allowing self-crossing. But at this time, the ambient temperature was already high and was not conducive to the germination of pollen grains. The corolla closed, forming an inverted bell shape, where the inner microenvironment ensured completion of pollen germination and fertilization. Preferential cross-pollination and delayed self-crossing of I. purpurea provided a doubly guaranteed mechanism for pollination and fertilization, facilitating its adaptation to a high temperature climate.

[Analysis of mental state of allergic rhinitis patients in Chengdu city by symptom check list 90 (SCL-90) scale].

Objective: To analyse the mental state of patients with allergic rhinitis (AR) in Chengdu. Methods: One thousand five hundred and thirty-six AR patients from Sichuan Provincial Integrated Traditional Chinese and Western Medicine Hospital, West China Hospital of Sichuan University, Affiliated Hospital of Chengdu University of Traditional Chinese Medicine, Sichuan People's Hospital, Sichuan Second Hospital of Traditional Chinese Medicine were selected from July 2013 to January 2018. Eight hundred and twenty-seven patients were screened into study group by inclusion and exclusion standards. The symptom check list 90 (SCL-90) was used to group and score the mental state of these patients according to nine classification criteria: gender, BMI, age, marital status, monthly salary, disease duration, living environment, education level and working environment. Then, the scores were compared within groups. Inter-group comparison was made between the study group and the Chinese norm, and the positive factors for psychological disorders were extracted. Four symptoms in the study group, i.e. nasal itching, sneezing, clear discharge and nasal congestion, were scored on the visual analogue scale (VAS). SPSS 19.0 software was used to carry out statistical analysis. Partial correlation analysis was performed between the positive factors and the symptom scores by multiple regression statistical method. Results: The total score of SCL-90 in the study group was 2.64±0.25, which was accorded with mild to moderate mental health impairment. There were 124 (15.0%) without mental health damage, 176 (21.3%) with mild damage, 474 (57.3%) with mild to moderate damage, 41 (5.0%) with moderate to severe damage and 12 (1.4%) with severe damage. The in-group comparison showed that the top three categories of different items were the living environment, gender and working environment. The scores of somatization, obsessive-compulsive symptoms, interpersonal sensitivity, depression, anxiety, psychosis, other (sleep, diet) and total average score of urban residents were higher than that of country residents (3.29±0.61 vs 2.65±0.50, 2.81±0.77 vs 2.05±0.38, 3.10±0.19 vs 2.49±0.67, 3.40±0.84 vs 2.49±0.70, 3.04±0.64 vs 2.33±0.51, 3.02±0.55 vs 2.40±0.77, 3.40±0.41 vs 2.52±0.77, 2.91±0.11 vs 2.29±0.40, Z value was 4.88, 5.25, 4.57, 5.91, 5.09, 4.63, 5.55, -4.55, respectively, all P<0.05). Women scored higher than man for somatization, interpersonal sensitivity, depression and others (2.66±0.51 vs 2.00±0.45, 3.37±0.47 vs 2.63±0.51, 3.44±0.57 vs 2.85±0.52, 3.47±0.36 vs 2.76±0.45, Z value was -5.10, -5.51, -4.86, -5.28, respectively, all P<0.05). The scores of somatization, interpersonal sensitivity, psychosis and other (sleep, diet) were higher in the indoor group than those in the outdoor group (3.49±0.64 vs 2.78±0.46, 3.33±0.30 vs 2.56±0.68, 3.28±0.60 vs 2.67±0.31, 3.50±0.85 vs 2.85±0.37, Z value was 5.31, 5.79, 4.89, 5.00, respectively, all P<0.05). The outdoor group scored higher on obsessive-compulsive symptoms, anxiety and hostility (3.44±0.40 vs 2.83±0.35, 3.40±0.50 vs 2.57±0.93, 3.34±0.88 vs 2.69±0.56, Z value was 4.96, 6.22, 5.08, respectively, all P<0.05). The inter-group comparison found that depression, anxiety, psychosis and other (sleep, diet) could be partially correlated with VAS scores as 4 positive factors. The results of partial correlation analysis showed that depression was positively correlated with sneezing and nasal runny discharge, anxiety was positively correlated with nasal itching and nasal obstruction, psychosis was positively correlated with nasal itching and sneezing, and other (sleep, diet) was positively correlated with nasal runny discharge and nasal obstruction. Conclusion: AR patients have mild to moderate mental health impairments, which are correlated with AR symptoms.

Melatonin regulates mitochondrial function and biogenesis during rat dental papilla cell differentiation.

OBJECTIVE: The aim of this study was to investigate the effect of melatonin on mitochondria of dental papilla cells (DPCs) during the odontogenic differentiation process. MATERIALS AND METHODS: Primary DPCs were obtained from the first molar dental papilla of neonatal rats and cultured in osteogenic (OS) or basal medium supplemented with melatonin at different concentrations (0, 1 pM, 0.1 nM, 10 nM, and 1 µM) for differentiation in vitro. Effects of melatonin on differentiation, mitochondrial respiratory function, and mitochondrial biogenesis of DPCs were analyzed. RESULTS: Upon odontogenic induction, Alkaline phosphatase (ALP) activity, dentin sialophosphoprotein (DSPP), and dentin matrix protein (DMP1) expression were significantly enhanced, with a peaked expression at 10 nM of melatonin treatment. During DPCs differentiation, 10 nM melatonin could significantly induce the increase of intracellular Adenosine triphosphate (ATP), the decrease of the oxidized form of nicotinamide adenine dinucleotide (NAD+)/NADH ratio and reactive oxygen species (ROS). The mRNA and protein levels of peroxisome proliferator-activated receptor gamma coactivator 1-alpha (PGC-1α), nuclear respiratory factor 1 (NRF-1), and mitochondrial transcription factor A (TFAM) were significantly increased, and the peak level of expression was found in cells treated with 10 nM of melatonin. Furthermore, the mitochondria DNA (mtDNA) copy number was significantly decreased during DPCs differentiation. CONCLUSIONS: These findings suggest that melatonin can promote the differentiation of rat DPCs and regulate mitochondrial energy metabolism, ROS scavenging, and mitochondrial biogenesis.

Gene-based GWAS analysis for consecutive studies of GEFOS.

By integrating the multilevel biological evidence and bioinformatics analyses, the present study represents a systemic endeavor to identify BMD-associated genes and their roles in skeletal metabolism. INTRODUCTION: Single-nucleotide polymorphism (SNP)-based genome-wide association studies (GWASs) have already identified about 100 loci associated with bone mineral density (BMD), but these loci only explain a small proportion of heritability to osteoporosis risk. In the present study, we performed a gene-based analysis of the largest GWASs in the bone field to identify additional BMD-associated genes. METHODS: BMD-associated genes were identified by combining the summary statistic P values of SNPs across individual genes in the two consecutive meta-analyses of GWASs from the Genetic Factors for Osteoporosis (GEFOS) studies. The potential functionality of these genes to bone was partially assessed by differential gene expression analysis. Additionally, the consistency of the identification of potential bone mineral density (BMD)-associated variants were evaluated by estimating the correlation of the P values of the same single-nucleotide polymorphisms (SNPs)/genes between the two consecutive Genetic Factors for Osteoporosis Studies (GEFOS) with largely overlapping samples. RESULTS: Compared to the SNP-based analysis, the gene-based strategy identified additional BMD-associated genes with genome-wide significance and increased their mutual replication between the two GEFOS datasets. Among these BMD-associated genes, three novel genes (UBTF, AAAS, and C11orf58) were partially validated at the gene expression level. The correlation analysis presented a moderately high between-study consistency of potential BMD-associated variants. CONCLUSIONS: Gene-based analysis as a supplementary strategy to SNP-based genome-wide association studies, when applied here, is shown that it helped identify some novel BMD-associated genes. In addition to its empirically increased statistical power, gene-based analysis also provides a higher testing stability for identification of BMD genes.

[Therapeutic effect of cervical Jiaji electroacupuncture on postoperative intractable hiccup of liver neoplasms].

Objective: To evaluate the therapeutic effect of cervical Jiaji electroacupuncture on postoperative intractable hiccup of liver neoplasms. Methods: A total of 39 patients with postoperative intractable hiccup of liver neoplasms in The First Affiliated Hospital of Heilongjiang University of Chinese Medicine from May 2013 to May 2017 were collected and divided into 2 groups randomly. The electroacupuncture group included 20 cases, the control group included 19 cases. Patients in the electroacupuncture group were treated by cervical Jiaji electroacupuncture (located in C3-5, sympathetic ganglion), while the control group were treated by metoclopramide combined with chlorpromazine for three days. The therapeutic effects of two groups were compared and the onset time were recorded. Results: Total effective rates of electroacupuncture group and control group were 95.0% and 47.4%, respectively. The onset time in electroacupuncture group and control group were (14.8±3.3) h and (30.5±3.1) h, respectively (P<0.01). Ten cases who resisted the control treatment were then treated by electroacupuncture for 3 days, 6 cases were recovered, 3 cases became better, while 1 case demonstrated no response. No serious adverse reactions were appeared in each group. Conclusion: Cervical Jiaji electroacupuncture is an effective and safe treatment for postoperative intractable hiccup of liver neoplasms, and it can be used as a remedy for intractable hiccup patients who don't respond to drug treatment.

First Report of Black Spot of Acanthus ilicifolius Caused by Fusarium solani in China.

Acanthus ilicifolius (family Acanthaceae) grows mainly in tropical coastal areas and is an important medicinal plant that can be used to treat asthma, rheumatism, etc. In July 2013, symptoms of black spots on the leaves of A. ilicifolius were observed in the Mangrove Conservation Area of Shenzhen Futian (22°32' N, 114°03' E) and Leizhou peninsula (20°12'~21°35' N, 109°30'~110°55' E), Guangdong Province, China. Initial symptoms of the disease were a small, dark brown spots (4 to 5 × 4 to 6 mm) surrounded by a yellow halo (1 to 2 mm in diameter), that would later extend to round or irregular black spots. Leaves eventually turned chlorotic and plants defoliated. Tissues from symptomatic leaves were excised, surface sterilized with 75% ethanol solution (v/v) for 20 s, soaked in 0.1% HgCl2 solution for 45 s, rinsed three times in sterile water, cut into small pieces (2 to 3 mm), plated on potato dextrose agar (PDA), and incubated 3 to 5 days at 28°C without light. Four isolates named from LSL-1 to LSL-4 with different morphological characteristics were obtained. To fulfill Koch's postulates, wounded and non-wounded leaves were inoculated. Fresh wounds were made with a sterile needle on detached leaves and on living plants. Mycelial plugs of each isolate were applied and covered with a piece of wet cotton to maintain moisture. For the control, the healthy leaves were inoculated with PDA plugs. All treatments were incubated at room temperature. Black spots were observed on the wounded leaves inoculated with isolate LSL-1 after 3 days, while the other three isolates and the control remained symptomless, and the pathogen similar to LSL-1 was re-isolated from the diseased leaves. Non-wounded leaves didn't become infected. The pathogenic test was repeated three times with the same conditions, and it was confirmed that LSL-1 was the pathogen causing the black spot of A. ilicifolius. Identification of the pathogen was conducted using morphological and molecular characteristics. Hyphal tips of LSL-1 were transferred to PDA medium in petri dishes for morphological observation. Two types of conidia were observed. The macroconidia were cylindrical to slightly curved, falciform shaped, with two to four septa, and measured 39 to 45 × 4.7 to 5.0 µm. The microconidia were oval to kidney shaped, single celled, 8 to 10 × 2.5 to 3.5 µm. Chlamydospores were also observed, produced singly or in pairs. Based on morphology (1,4), the isolate was tentatively identified as Fusarium solani. For molecular identification, the internal transcribed spacer (ITS) of ribosomal DNA, beta-tubulin gene, and translation elongation factor 1-alpha (EF-1α) gene was amplified using the ITS1/ITS4 (5), ITS4/ITS5 (5), T1/T2 (2) and EF1/EF2 (3) primer pairs. The gene sequences were deposited in GenBank (KJ720639 for the ITS1/ITS4 region, KF826493 for the ITS4/ITS5 region, KJ720638 for the beta-tubulin, and KF826492 for EF-1α region) and showed 99% identity to the F. solani strains (AY633746 for ITS1/ITS4 region, AM412637 for ITS4/ITS5 region, KF255996 for beta-tubulin region, DQ246859 for EF-1α region). According to these results, the pathogen of black spot of A. ilicifolius was identified as F. solani. To the best of our knowledge, this is the first report of F. solani causing black spot of A. ilicifolius in China. References: (1) J. F. Leslie and B. A. Summerell. The Fusarium Laboratory Manual. Blackwell, Ames, IA, 2006. (2) K. O'Donnell and E. Cigelnik. Mol. Phylogenet. Evol. 7:103, 1997. (3) K. O'Donnell et al. Proc. Natl. Acad. Sci. USA. 95:2044, 1998. (4) B. A. Pérez et al. Plant Dis. 91:1053, 2007. (5) A. W. Zhang et al. Plant Dis. 81:1143, 1997.

Relationship between the antibacterial and immunological activities of houttuyfonate homologues and their surface activities.

Based on the microbial model (Staphylococcus aureus and Bacillus subtilis) and the experiment of mouse carbon granular clearance, the relationship between antibacterial and immunological activities of houttuyfonate (HOU) homologues and their surface activities were studied. The results showed that with elongating the carbon chain of HOU homologues, the surface activities and bacteriostasis and immunological activities would be increased. It is suggested that the pharmacological effect of HOU homologues could be realized by the hydrophobic interaction between HOU homologues and membrane proteins of bacteria and cell.

[Study on the tissue culture and plant regeneration of Pogostemon cablin].

Root tips, leaf segments, stem segments with node and stem segments of Pogostemon cablin (Blanco) Benth. were cultured in vitro. The results indicated that leaf segments and stem segments with node were suitable for callus formation, the frequency of callus induction were over 87.0%. A optimal medium for callus induction was MT + BA 0.5 mg/L + NAA 0.2 mg/L. MT basal medium with BA 0.5-1.0 mg/L were effective to induce differentiation of callus. And at the MT basal medium, the regenerative shoots from Pogostemon cablin could form roots.

Location and migration of cations in Cu(2+)-adsorbed montmorillonite.

Locations of Cu2+ ion in Cu(2+)-adsorbed montmorillonite have been studied by electron paramagnetic resonance (EPR), supplemented by X-ray diffraction (XRD) and differential thermal analysis (DTA). In the EPR spectra of Cu(2+)-adsorbed montmorillonite, three signals, corresponding to Cu2+ ion, have been simultaneously recorded. Some Cu2+ ions seemed to replace the original interlayer metal cations and some entered into the hexagonal cavities. A small fraction of Cu2+ ions penetrated into the octahedral vacancies. There were two ways for the adsorption of Cu2+ ion by montmorillonite--exchangeable and specific. On heating, the hydrated Cu2+ ion in the interlayer loses the coordinating water and then enters into the hexagonal cavities. When the heating temperature further increased, dehydroxylation occurs, which facilitates Cu2+ ion in the hexagonal cavities to penetrate into the octahedral vacancies.

New diterpene alkaloids from the roots of Spiraea japonica.

Chemical investigation on an ethanol extract from the roots of Spiraea japonica var. acuta resulted in the isolation of three new diterpene alkaloids named spiramide (1) and spiratine A (2) and spiratine B (3). Structures of 1-3 were elucidated primarily on the basis of 1D and 2D NMR experiments.

[Plant regeneration from Agobacterium-mediated CTV-cp gene transformation of Poncirus trifoliata Raf].

OBJECTIVE: To lay a foundation for the resistant breeding, the anti-virus CTV-cp gene was transformed into the epicotyles mediated by Agrobacterium tumefaciens in Poncirus trifoliata. METHOD: The explants used for the genetic transformation were the epicotyls from P. trifoliata. The Agrobacterum tumefaciens strain was EHA101 containing vector plasmid pGA482GG. The coat protein gene (CTV-cp gene), GUS gene and NPT II gene were introduced into the transformation plasmid. RESULTS: Ceftaxime used as antibiotics was better than carbenicillin. The concentration was 300 mg.L-1; The selection pressure for kanamycin was 50 mg.L-1; 70.0% of the resistant plants were GUS-positive; extra gene was proved to be in P. trifoliata plant by southern blot examination. CONCLUSION: An effective genetic transformation mediated by Agrobacterium tumefacines, which harbours a CTV-cp gene, has been developed in P. trifoliata. Transgenic CTV-cp plants were obtained.

Cloning of human myeloid-associated differentiation marker (MYADM) gene whose expression was up-regulated in NB4 cells induced by all-trans retinoic acid.

A full-length cDNA of 3192 bp isolated from human bone marrow cDNA library was predicted an ORF encoding 298 amino acids. The deduced protein, containing seven putative transmembrane segments and sharing 75.8% amino acid identity with mouse Myadm protein, was named as human MYADM. The results of Northern blot analysis showed that MYADM was ubiquitously expressed in 15 of 16 adult tissues tested, except thymus. To determine whether the novel human gene was involved in hematopoietic differentiation process as mouse Myadm did, we examined the mRNA expressive abundance of this gene between normal bone marrow cells and peripheral blood leukocytes, and detected the expression change in NB4 cells induced by all-trans retinoic acid at different induce time by the semi-quantitative RT-PCR. The results showed that the expression of the novel gene was not only significantly higher in peripheral blood leukocytes than in bone marrow cells, but also significantly up-regulated when the NB4 cells(derived from a patient with acute promyelocytic leukemia) were induced by all-trans retinoic acid (ATRA) for 48hr. It is suggested that human MYADM was also associated with the differentiation of hematopoietic cells or acute promyelocytic leukemia cells. In addition, MYADM was mapped to human chromosome 19q 13.33-q 13.4 by Radiation Hybrid mapping, and it consists of 3 exons and 2 introns and spans a 7.1-Kb genomic region.

Prenatal morphine exposure differentially alters TH-immunoreactivity in the stress-sensitive brain circuitry of adult male and female rats.

Previously, we demonstrated that exposure to morphine during gestation increases hypothalamic norepinephrine (NE) content and turnover rate in adult male rats and decreases these measures in adult females. To investigate the basis of these alterations, the present study examined the effects of prenatal exposure to morphine on tyrosine hydroxylase immunoreactivity (TH-IR) in the brains of adult male and female progeny. In male rats, prenatal morphine exposure significantly increased the density of TH-IR in cells and fibers in the caudal paraventricular nucleus of the hypothalamus (PVN) and locus coeruleus (LC), but had no effects in the lateral hypothalamus (LH). In female rats that were ovariectomized (OVX), prenatal morphine exposure significantly decreased the density of TH-IR in cells and fibers in the LC. Interestingly, an injection of estrogen in OVX control females reduced the mean optical density of TH-IR in the LC, but it was ineffective in drug-exposed females in the same brain region. Estrogen injections also reduced the mean optical density of TH-IR in the LH but not in the PVN of females, regardless of prenatal drug exposure. Thus, the present study suggests that prenatal morphine exposure induces long-term, sex-specific alterations in TH-IR in the PVN and LC of adult progeny.

[Effect of different watering and fertilization on N utilization efficiency].

The comprehensive effects of the timing of watering and N and K fertilization(D), amount of irrigated water(W), and application rates of chemical N and K fertilizers, and farm manure(M) were studied by the design of orthogonal regression rotation with five factors and five levels under dry greenhouse condition. The N utilization efficiency of potato was in order of N > W > D > M > K. Under different application rates of W, N, K and M, the N utilization efficiency was significantly increased with optimum timing of watering and fertilization. When the application rate was low, the timing should be earlier, while it was high, the timing should be later. High application rates of W and N resulted in the best interaction effect, with 57.83% of N utilization efficiency; moderater W and N or low W and N had 29.17-40.99% of efficiency; and high W and low N or low W and high N resulted in the lowest efficiency, with 22.87% and 22.51%, respectively.