italic>α-Conotoxin ArIB[V11L,V16D] is currently the most optimal selective inhibitor of α7 nicotinic acetylcholine receptor (nAChR) known. In order to explore chemical modification methods and enrich its application in targeting nAChR, this study utilized the linker to covalently connect camptothecin and 7-amino-4-methylcoumarin to the [2,4] disulfide bond of ArIB[V11L,V16D]. Therefore, two peptide-drug conjugates (PDCs), ArIB[V11L,V16D]-5 and ArIB[V11L,V16D]-6, and one fluorescent-labeled peptide, ArIB[V11L,V16D]-7 were constructed. Cytotoxicity evaluation showed that the IC₅₀ values against non-small cell lung cancer cell line A549 of the two PDCs were respectively 1.3 and 4.1 times of camptothecin, indicating slight reduction in activity at the cellular level which was related to the linker structure. Fluorescence spectrum scanning revealed that the excitation and emission wavelength of the fluorescent-labeled peptide were 340 nm and 403 nm respectively, and the fluorescence features of 7-amino-4-methylcoumarin as a marker were retained without fluorescence quenching. This modification strategy laid a solid foundation for the further application of α-conotoxin ArIB[V11L,V16D] in PDCs and fluorescent probes.

Objective To investigate the prevalence,genotype and epidemiology of plasmid- mediated AmpC enzyme of Escherichia coli and Klebsiella pneumoniae.Methods A total of 67 clinical isolates of nonrepetitive cefoxitin-resistant Escherichia coli and Klebsiella pneumoniae collected by Fuzhou General Hospital and Fujian Provincial Hospital during a period of Sept.2004 to Mar.2005 were detected by three-dimensional extract test for AmpC enzyme,and PCR for AmpC enzyme and other ?-lactamase gene amplification and DNA sequencing were carried out for genotype of ?-lactamase.Plasmid transformation experiment was used to study the transfer of cefoxitin resistance.The homology of the isolates was determined by ERIC-PCR fingerprinting.Results At two hospitals in Fuzhou,the prevalence of plasmid-mediated AmpC enzyme among cefoxitin-resistant Escherichia coli and Klebsiella pneumoniae were 16.7% and 10.5%, 8.0% and 0,respectively.Two isolates of Klebsiella pneumoniae produced DHA-1 plasmid-mediated AmpC enzyme,and 4 isolates of Escherichia cob and one strain of Escherichia coli produced CMY-2 and CMY-22 plasmid-mediated AmpC enzyme respectively.Furthermore,5 strains of Escherichia coli with CMY AmpC enzyme were also found simuhaneously to produce TEM-144,CTX-M-27,CTX-M-14 and TEM-1 ?-lactamase respectively.Three strains of Escherichia coli and one isolate of Klebsiella pneumoniae could transfer cefoxitin resistance to acceptant bacillus.ERIC-PCR fingerprinting reveals 2 strains of Klebsiella pneumoniae came from same clone,but 5 strains of Escherichia coli came from different clones.Conclusions The clinical isolates of Klebsiella pneumoniae producing DHA-1 plasmid-mediated AmpC enzyme and Escherichia coli producing CMY-2,CMY-22 plasmid-mediated AmpC enzyme are found in Fuzhou.CMY-22 AmpC enzyme and TEM-144 ?-lactamase are the first reported in the world,GenBank accession number: DO256079,DO256080

ObjectiveTo find out if the immune system derived thyroid stimulating hormone(TSH) β splice variant(TSHβ-Ⅴ) would be regulated by circulating thyroid hormone levels to get a further understanding of the function and mechanism of this TSHβ-Ⅴ in thyroid homeostasis.MethodsA total of 20 weaning Balb/c mice (half male and half female) were selected and randomly divided into two groups according to their body mass and gender(n =10).Mice of control group were fed with common diet and deionized water.Mice of the low-iodine(LI) group were fed with low-iodine diet(containing iodine 20 - 40 μg/kg,iodine-intake about 0.25 μg/d) and deionized water.The experimental period was 3 months.At the end of the experiment,mice were executed and the blood was collected to observe the levels of TSH and thyroid hormone by chemiluminescence immunoassay (CIA) ; bone marrow (BM),peripheral blood(PBL),thyroid gland and pituitary were collected to assay the TSHβ-Ⅴ mRNA expression by real-time quantitative reverse transcriptase polymerase chain reaction (RT-PCR).ResultsThe serum free thyroxine(FT4) and total thyroxine(TT4) levels in LI group of mice[(0.47 ± 0.70)nmol/L,(2.41 ± 0.28)pmol/L] were significantly lower than that of the control group of mice [(55.2 ± 3.68) nmol/L, (32.72 ± 1.02) pmol/L,t =43.81,86.04 、all P ＜ 0.01 ] and the serum total triiodothyronine(TT3) and free triiodothyronine(FT3) reduction in LI group of mice[ (0.76 ± 0.08)nmol/L,(4.01 ± 0.40)pmol/L] were significantly lower than that of the control group of mice [ (1.10 ± 0.06)nmol/L,(5.40 ± 0.38)pmol/L,t =9.81,7.5 1,P ＜ 0.01 ].Iodine insufficiency strongly elevated the serum TSH in LI group of mice[ (35.67 ± 17.39)mU/L] than that in control group of mice[ (0.24 ± 0.10)mU/L,t =- 6.11,P ＜ 0.01 ].The mRNA levels of TSH β-Ⅴ in BM (9.62 ± 0.60) and in PBL( 9.25 ± 0.83 ) of LI group of mice were lower than those in control group of mice (7.69 ± 0.36,7.11 ± 0.41,t =6.77,5.64,P ＜ 0.01),while the mRNA level of TSH β-Ⅴ in pituitary of LI group of mice (1.99 ± 0.61) was increased compared with that in control group of mice (5.75 ± 0.98,t =- 8.02,P＜ 0.01).Compared with control group of mice(9.12 ± 0.62),the level of thyroid TSH β-Ⅴ mRNA in LI group of mice (9.32 ± 0.91 ) was not significantly changed (t =0.45,P ＞ 0.05).There was no detectable native TSHβ in BM,PBL and thyroid.The mRNA level of native TSHβ in pituitary in LI group of mice( - 7.17 ± 1.78) was dramatically elevated compared to that in control group of mice( - 1.43 ± 0.51,t =- 7.60,P ＜ 0.01 ).ConclusionsThe mRNA levels of TSHβ-Ⅴ are suppressed in BM and PBL in low iodinediet induced hypothyroidism mice,which suggest that immune system derived TSHβ-Ⅴ may be more important thannative TSHβ in immune-thyroid regulation.

Objective To detect the incidence of inherited metabolic diseases(IMD)and disorders of metabo-lism in 4 710 high - risk infants,as well as providing basis of clinical diagnosis and treatment by using urease pretreat-ment - gas chromatography - mass spectrometry(UP - GC - MS). Methods Samples were collected from high - risk infants with IMD,after removing urea,putting in internal standard,removing protein,vacuum drying and bis (trimethyl - silyl)trifluoroacetamide / trimethyl - chlorosilane derivativing,UP - GC - MS was used to analyze compo-sitions such as organic acids,amino acids,carbohydrates,pyridoxines,purines and pyrimidines,then metabolic analysis was proceeded to refer to the normal detection value of the healthy children,finally a metabolic diagnosis was made ba-sing on the clinical data such as the high - risk clinical manifestations and general biochemical tests and other special examinations. Results In the 4 710 cases,there were 98 cases of IMD(2. 1% ),326 cases of suspected IMD(6. 9% ), 2 610 cases of metabolic disorders(55. 4% ). There were 98 cases of IMD,including 57 cases of methylmalonic aciduria,12 cases of propionic acidemia,7 cases of glutaric aciduria,5 cases of hyperphenylalaninemia,maple syrup u-rine disease and multiple carboxylase defects each,4 cases of isovaleric acidemia and 3 cases of 4 - hydroxy butyric acid urine disease. Conclusions UP - GC - MS is a effective way to diagnose IMD and metabolic disorders of infants. Common IMD in Guangdong Province include methylmalonic aciduria,propionic academia,glutaric aciduria,hyperphe-nylalaninemia,maple syrup urine disease and multiple carboxylase defects. The results of the tests can provide effective guidance for diagnosis and treatment of suspected infants.

Objective To observe and measure the anatomical structure of approach of vertical facial nerve canal and put forward the normal measurement range and the location relationship among the vertical segment of facial nerve canal, the posterior wall of external acoustic meatus and the rear edge of external ear,and discuss the relationship and clinical significance between the mastoid gasification and the vertical segment of facial nerve canal. Methods 1. Evaluate the accuracy of CT image of related structure, using spiral CT in scanning four skull specimens, get the horizontal distances of the vertical segment of facial nerve canal to the posterior wall of external acoustic meatus and the rear edge of external in the axial position, and get the sagittal diameter of mastoid (the horizontal distance from the lowest point of external auditory inferior canal to the rear edge of mastoid) and the height (the vertical distance between the lowest points of the external auditory canal wall to the mastoid tip) in the sagittal position. And then saw the skull specimens to measure the distance in the same lay with CT image, and discuss the statistics difference of the distance between the values of CT imaging measurements and the dry entities cranial measurements on hand. 2. Study on people: 118 patients (236 sides) with non-ear disorders were randomly selected, among which there were 63 females (126 sides) and 55 males (110 sides). They were subjected to maxillofacial CT scan in the same layer that used above, and the horizontal distances of the facial nerve canal to the posterior wall of external acoustic meatus and the rear edge of external ear were measured. In addition, half of the product of diameter and height of the mastoid was defined as mastoid area, which was used to define the extension of mastoid gasification. Then related analysis and regression analysis were done between the vertical segment of facial nerve canal and the posterior wall of external acoustic meatus, as well as the rear edge of external ear. Results 1.Part of the experiment: There was no significantly different on the indicator values between CT image the entity measurements among the four skull specimens (P>0.05). 2. Study on people: There was no significantly different between left side and right side(P>0.05), but significantly different between genders(P<0.05). Between mastoid area and the distance from the vertical segment of facial nerve canal to the posterior wall of external acoustic meatus there is inverse correlation, and the relevance has the remarkable significance. However, there was no correlation between mastoid area and the vertical segment of facial nerve canal to the rear edge of external ear. Conclusion There was some relationship between the location of the vertical segment of facial nerve canal and external acoustic meatus. Anatomic position of vertical facial nerve cancal and the posterior wall of external acoustic meatus can be showed clearly. CT and in combination with primitive axial images may provide reliable evidence for the diagnosis facial nerve dieases and the choice of ear surgery route.

Objective To evaluate clinical efficiency and quality-of-life outcomes in treatment of severe pelvic organ prolapse by the Xiehe pelvic floor reconstruction surgery. Methods From Jun. 2006 to Dec. 2008, 277 severe pelvic organ prolapse patients with stage Ⅲ to Ⅳ from 8 hospitals in China were enrolled in this prospective study. Pelvic organ prolapse quantitative examination (POP-Q) and anatomic improvement in these patients after surgery were analyzed in this interim study. Comparisons of pelvic floor impact questionnaire-short form 7 (PFIQ-7) and pelvic floor distress inventory-short form 20 (PFDI-20) in these patients before and after surgery was used to evaluate quality of life. Comparison of pelvic organ prolapse-urinary incontinence sexual questionnaire (PISQ) in these patients before and after surgery was used to evaluate quality of sexual life. Results With a median follow-up of 14. 0 months (6 -28 months),twenty-three patients showed recurrent prolapse (8. 3%, 23/277), and anatomical success ( ＜ stage 2 in the treated compartment) was 91.7% (254/277). In this series, mesh exposure or erosion rate was 6. 9% (19/277). The postoperative de novo stress incontinence rate was 6. 5% (18/277). The scores for PFIQ-7 and PFDI-20, and its subscales were significantly improved, the scores of before treatment were lower than those after treatment (P ＜0. 01 ). And there was no significant difference in the average score of PISQ before and after the surgery (76. 6 ± 15.4 versus 75.5 ± 14. 5 versus 73.6 ± 12. 6, P ＞0. 05 ), but the rate of de novo dyspareunia was 11% (9/80). Conclusions Xiehe pelvic floor reconstruction surgery was safe and efficacy in treatment of pelvic organ prolapse. It could improve quality of life remarkably with less cost when compared with the traditional total pelvic floor reconstruction surgery.

The traditional Chinese medicine of Radix Hedysari plays an important role in invigorating gas for as-cending, benefiting blood for promoting production of fluid, and promoting circulation for removing obstruction in collaterals, which is consistent with the principle of treatment for osteoporosis. This study is designed to investigate the bioactive components on increasing peak bone mass (PBM) by exploring the spectrum-effect relationship between chromatography fingerprints and effect. Multiple indicators are selected to evaluate the pharmacological activity. In fingerprints, 21 common peaks are obtained, five of which are identified. Furthermore, gray relational analysis (GRA) is a quantitative method of gray system theory and is used to describe the correlation degree of common peaks and pharmacological activities with relational value. 21 components are then divided into three different regions, of which ononin and calycosin play an extremely significant role in increasing PBM. In addition, factor analysis and hierarchical cluster analysis (HCA) are used to screen the optimal producing area for Radix Hedysari. This provides a comprehensive and efficient method to improve the quality evaluation of Radix Hedysari, confirming the bioactive components for PBM-enhancement and further develop its medicinal value.

The direct acting substances of Cuscuta chinensis in vivo were preliminarily identified through the correlation analysis of "metabolites-effect identification" model. The ovariectomized female rats were i.g administered with 95% ethanol extract part, 40% ethanol elution part and n-butanol extract part of Cuscuta chinensis. The serum fingerprints of different parts and times of administration were established by UPLC/Q-TOF-MS. At the same time, serum estradiol (E₂), follicle-stimulating hormone (FSH) and luteinizing hormone (LH) levels were detected. Bivariate correlation analysis and grey correlation analysis were used to screen estrogenic components. The results showed that nine direct acting substances in vivo highly related to estrogen effect were found in the drug containing serum, which were hyperoside, astragalin, methyl quercetin glucuronide, quercetin-diglucuronide, quercetin, apigenin, isoquercitrin, kaempferol glucuronide and kaempferol. We can preliminarily screen out the direct acting substance of estrogen effect of Cuscuta chinensis in vivo based on the research idea of serum spectrum effect correlation. It provides a reliable basis for revealing the estrogeneffective substances of Cuscuta chinensis and confirming the quality markers. This experiment was approved by Harbin University of Commerce Ethics Committees (Approval No. HSDU2020-065).

OBJECTIVE: To establish a method of preparing and analyzing DNA of sperm cells from sample mixtures. METHODS: Sperm cells were captured by micromanipulation method, DNA template was prepared from these sperm cells, and then amplified by multiple displacement amplification (MDA). RESULTS: High-yield of highly integrated DNA was produced by MDA from very small amount of DNA template of 20 sperm cells. The DNA could be amplified at least 30,000-fold, and the length of most the DNA segments were greater than 15 kb, providing sufficient amount of short tandem repeat (STR) products for DNA genotyping. CONCLUSION The micromanipulation method is efficient to isolate sperm cells from sample mixtures and multiple displacement amplification can provide sufficient amount of DNA products for STR genotyping. The method seems to be feasible in detecting DNA from rape cases.
DNA/isolation & purification* ; Forensic Medicine/methods* ; Humans ; Male ; Nucleic Acid Amplification Techniques ; Spermatozoa/chemistry* ; Tandem Repeat Sequences

BACKGROUNDDisequilibrium of Th1/Th2 is known as an important cause of allergic asthma with a biased Th2 type response. It has been shown that lipopolysaccharide (LPS) administration during post-sensitization modified the inflammation of asthma via upregulating the Th1 response that decrease the Th2 immunity. We would like to know if, during pre-sensitization, the elevated Th1 response is necessary for LPS exposure to modify the asthmatic response.

METHODSDuring pre- or post-sensitization, 40 microg LPS were intraperitoneal injected (i.p.) to asthmatic mice sensitized and challenged by Dermatophagoides farinae (D. farinea). Inflammation was assessed by examining bronchoalveolar lavage fluid (BALF) for the number and identity of cells and by cytokine titers measured by ELISA. Semi-quantified RT-PCR was used to evaluate the level of Toll-like receptor 4 (TLR4) mRNA in dendritic cells (DCs) from bone marrow (BMDCs).

RESULTSThese investigations demonstrated that LPS exposure during pre-sensitization inhibited the Th2 cytokine and inflammatory infiltration, the same as with LPS exposure during post-sensitization in allergic asthma mice. Contrary to post-sensitization LPS exposure, the Th1 cytokines were not upregulated by pre-sensitization with LPS. Finally, the study failed to show any significant difference between TLR4 mRNA expressed in BMDCs with the two times of LPS exposure.

CONCLUSIONSOur data suggest that elevated Th1 immunity is not required for the modification of the Th2 response induced by LPS exposure during pre-sensitization in asthmatic mice and that pre-sensitization differs from post-sensitization. Immune modulation with treatment is independent of TLR4 expression in BMDCs. This study implicates a potential way to protect from allergic disease and an inflammatory response.

Animals ; Asthma ; chemically induced ; immunology ; Bronchoalveolar Lavage Fluid ; immunology ; Cytokines ; immunology ; metabolism ; Dendritic Cells ; immunology ; Dermatophagoides farinae ; immunology ; Female ; Lipopolysaccharides ; immunology ; Mice ; Mice, Inbred BALB C ; Reverse Transcriptase Polymerase Chain Reaction ; Th1 Cells ; immunology ; Th2 Cells ; immunology ; Toll-Like Receptor 4 ; genetics