A synthetic standard for competitive RT-PCR quantitation of 13 GABA receptor type A subunit mRNAs in rats and mice

1998 ◽  
Vol 85 (1) ◽  
pp. 89-98 ◽  
Author(s):  
Zhi Fang Liu ◽  
David R Burt
Author(s):  
David A. Nielsen ◽  
Dmitri Proudnikov ◽  
Mary Jeanne Kreek

Impulsivity is a complex trait that varies across healthy individuals, although when excessive, it is generally regarded as dysfunctional. Impulsive behavior may lead to initiation of drug addiction that interferes with inhibitory controls, which may in turn result in facilitation of the individual’s impulsive acts. Although environmental factors play a considerable role in impulsive behavior, a body of evidence collected in twin studies suggests that about 45% of the variance in impulsivity is accounted for by genetic factors. Genetic variants studied in association with impulsivity include those fortryptophan hydroxylase 1 and 2 (TPH1 and TPH2), the serotonintransporter (SERT), serotonin receptors, and genes of the monoamine metabolism pathway (e.g., monoamine oxidase A, MAOA). Other systems may also play a role in these behaviors, such as the dopaminergic system (the dopamine receptors DRD2, DRD3, and DRD4, and the dopamine transporter, DAT), the catecholaminergic system (catechol-O-methyltransferase, COMT), and the GABAergic system (GABAreceptor subunit alpha-1, GABRA1; GABA receptor subunit alpha-6, GABRA6; and GABA receptor subunit beta-1, GABRB1). Taking into account involvement of the hypothalamic-pituitary-adrenal (HPA) axis, the number of candidate genes implicated in impulsivity may be increased significantly and, therefore, may go far beyond those of serotonergic and dopaminergic systems. For a number of years, our group has conducted studies of the association of genes involved in the modulation of the stress-responsive HPA axis and several neurotransmitter systems, all involved in the pathophysiology of anxiety and depressive disorders, impulse control and compulsive disorders, with drug addiction. These genes include those of the opioid system: the mu- and kappa-opioid receptors (OPRM1 and OPRK1) and the nociceptin/orphaninFQ receptor (OPRL1); the serotonergic system: TPH1 and TPH2 and the serotonin receptor 1B (5THR1B); the catecholamine system: COMT; the HPA axis: themelanocortin receptor type 2 (MC2R or adrenocorticotropic hormone, ACTHR); and the cannabinoid system: the cannabinoid receptor type 1 (CNR1). In this chapter we will focus on these findings.


2017 ◽  
Vol 33 (1) ◽  
pp. 8-15
Author(s):  
LR Barman ◽  
RD Sarker ◽  
BC Das ◽  
EH Chowdhury ◽  
PM Das ◽  
...  

A virological survey for avian influenza (AI) and Newcastle disease (ND) was conducted in two selected live bird markets (LBMs), namely Kaptan Bazar and Karwan Bazar in Dhaka city, Bangladesh from August 2011 to July 2012. A total of 513 dead chickens were collected. An immune-chromatographic rapid antigen test for Type A influenza virus and both conventional and real time RT-PCR were used for the detection and characterization of AI and ND viruses. All carcasses were first screened by the rapid antigen test kit and 93 were positive for Type A influenza virus. RT-PCR on a representative number of rapid antigen test positive samples (n = 24) confirmed the presence of Type A influenza virus and mostly H5 influenza virus (22 out of 24 tested samples). Influenza rapid test negative samples (n = 420) were subjected to routine necropsy. Heat stress, suffocation and physical injury were the most common cause of mortality (163 cases), followed by ND, suspected to be the cause of 85 deaths. On molecular investigation of these 85 samples, the presence of ND virus was confirmed in 59 and AI virus in 6; 15 were negative for both ND and AI viruses and 5 were unsuitable for investigation. Among the 59 ND confirmed cases 18 also contained AI virus. In summary, out of 513 carcasses 117 (22.81%) contained AI virus and 59 (11.50%) contained ND virus. Eighteen (3.51%) carcasses contained both AI and ND viruses. The findings suggest that both AI and ND should be considered as major threats to the poultry industry.Bangl. vet. 2016. Vol. 33, No. 1, 8-15


Author(s):  
Natalia L. Rukavina Mikusic ◽  
María I. Rosón ◽  
Nicolás M. Kouyoumdzian ◽  
Silvana M. Cantú ◽  
Belisario E. Fernández ◽  
...  

2017 ◽  
Vol 9 (1) ◽  
Author(s):  
Xin Wang ◽  
Shisong Fang

ObjectiveTo determine avian influenza A(H5N6) virus infection in humanand environment using extensive surveillances. To evaluate theprevalence of H5N6 infection among high risk population.IntroductionSince the emergence of avian influenza A(H7N9) virus in 2013,extensive surveillances have been established to monitor the humaninfection and environmental contamination with avian influenza virusin southern China. At the end of 2015, human infection with influenzaA(H5N6) virus was identified in Shenzhen for the first time throughthese surveillances. These surveillances include severe pneumoniascreening, influenza like illness (ILI) surveillance, follow-up onclose contact of the confirmed case, serological survey among poultryworkers, environment surveillance in poultry market.MethodsSevere pneumonia screening was carried out in all hospitals ofShenzhen. When a patient with severe pneumonia is suspected forinfection with avian influenza virus, after consultation with at leasttwo senior respiratory physicians from the designated expert paneland gaining their approval, the patient will be reported to local CDC,nasal and pharyngeal swabs will be collected and sent for detectionof H5N6 virus by RT-PCR.ILI surveillance was conducted in 11 sentinel hospitals, 5-20 ILIcases were sampled for detection of seasonal influenza virus by RT-PCR test every week for one sentinel. If swab sample is tested positivefor influenza type A and negative for subtypes of seasonal A(H3N2)and A(H1N1), it will be detected further for influenza A(H5N6) virus.Follow-up on close contacts was immediately carried out whenhuman case of infection with H5N6 was identified. All of closecontacts were requested to report any signs and symptoms of acuterespiratory illness for 10 days, nasal and pharyngeal swabs werecollected and tested for influenza A(H5N6) virus by RT-PCR test.In the meantime, environmental samples were collected in the marketwhich was epidemiologically associated with patient and tested forH5N6 virus by RT-PCR test.Serological survey among poultry workers was conducted in tendistricts of Shenzhen. Poultry workers were recruited in poultrymarkets and screened for any signs and symptoms of acute respiratoryillness, blood samples were collected to detect haemagglutination-inhibition (HI) antibody for influenza A(H5N6) virus.Environment surveillance was conducted twice a month in tendistricts of Shenzhen. For each district, 10 swab samples werecollected at a time. All environmental samples were tested forinfluenza A(H5N6) virus by RT-PCR test.ResultsFrom Nov 1, 2015 to May 31, 2016, 50 patients with severepneumonia were reported and detected for H5N6 virus, three patientswere confirmed to be infected with H5N6 virus. Case 1 was a 26 yearsold woman and identified on Dec 29, 2015. She purchased a duck ata live poultry stall of nearby market, cooked and ate the duck 4 daysbefore symptom onset. After admission to hospital on Dec 27, hercondition deteriorated rapidly, on Dec 30 she died. The case 2 was a25 years old man and confirmed on Jan 7, 2016. He visited a marketeveryday and had no close contact with poultry, except for passingby live poultry stalls. He recovered and was discharged from hospitalon Jan 22. The case 3 was is a 31 years old woman and reported onJan 16, 2016, she had no contact with live poultry and died on Feb 8.For 60 close contacts of three cases, none of them reported signsor symptoms of acute respiratory illness, all of nasal and pharyngealswabs were tested negative for influenza A(H5N6) virus by RT-PCRtest. Of 146 environmental swabs collected in the case’s living placesand relevant poultry markets, 38 were tested positive for influenzaA(H5N6) virus by RT-PCR test.From Nov 1, 2015 to May 31, 2016, 2812 ILI cases were sampledand tested for influenza type A and subtypes of seasonal influenza.Those samples tested positive for influenza type A could be furthersubtyped to seasonal A(H3N2) or A(H1N1), therefore no sample fromILI case was tested for influenza A(H5N6) virus.Serological surveys among poultry workers were conductedtwice, for the first survey 186 poultry workers were recruited in Oct2015, for the second survey 195 poultry workers were recruited inJan 2016. Blood sample were collected and tested for HI antibodyof influenza A(H5N6) virus. 2 individuals had H5N6 HI antibodytiter of 1:40, 5 individuals had H5N6 HI antibody titer of 1:20, rest ofthem had H5N6 HI antibody titer of <1:20. According to the WHOguideline, HI antibody titer of≥1:160 against avian influenza viruswere considered positive.From Nov 1, 2015 to May 31, 2016, of 1234 environmental swabscollected in poultry markets, 339 (27.5%)were tested positive forinfluenza A(H5N6) virus by RT-PCR test. Each of the ten districtshad poultry markets which was contaminated by influenza A(H5N6)virus.ConclusionsIn 2015-2016 winter, three cases of infection with influenzaA(H5N6) virus were identified in Shenzhen, all of them were youngindividuals with average age of 27.3 years and developed severepneumonia soon after illness onset, two cases died. For acute andsevere disease, early detection and treatment is the key measure forpatient’s prognosis.H5N6 virus was identified in poultry market and other placeswhere patient appeared, implying poultry market probably was thesource of infection. Despite the high contamination rate of H5N6virus in poultry market, we found that the infection with H5N6 virusamong poultry workers was not prevalent, with infection rate being0/381. Human infection with H5N6 virus seemed to be a sporadicoccurrence, poultry-human transmission of H5N6 virus might not bevery effective.


Blood ◽  
1998 ◽  
Vol 91 (6) ◽  
pp. 1882-1890
Author(s):  
David S. Viswanatha ◽  
I.-Ming Chen ◽  
Pu Paul Liu ◽  
Marilyn L. Slovak ◽  
Cathy Rankin ◽  
...  

The inv(16)(p13q22) and t(16;16)(p13;q22) cytogenetic abnormalities occur commonly in acute myeloid leukemia (AML), typically associated with French-American-British (FAB) AML-M4Eo subtype. Reverse transcriptase-polymerase chain reaction (RT-PCR) techniques have been recently developed to detect the presence of several variants of the resultant CBFB-MYH11 fusion gene that encodes a CBFβ-smooth muscle myosin heavy chain (SMMHC) fusion protein. We have now determined the clinical use of a polyclonal antibody [anti-inv(16) Ab] directed against a junctional epitope of the most common type of CBFβ-SMMHC fusion protein (type A), which is present in 90% of inv(16)/t(16;16) AML cases. Using flow cytometry, reproducible methods were developed for detection of CBFβ-SMMHC proteins in permeabilized cells; flow cytometric results were then correlated with cytogenetics and RT-PCR detection methods. In an analysis of 42 leukemia cases with various cytogenetic abnormalities and several normal controls, the anti-inv(16) Ab specifically detected all 23 cases that were cytogenetically positive for inv(16) or t(16;16), including a single AML case that was RT-PCR–negative. In addition to detecting all type A fusions, the anti-inv(16) Ab also unexpectedly identified the type C and type D CBFβ-SMMHC fusion proteins. Molecular characterization of one RT-PCR–positive and Ab-positive t(16;16) case with a non-type A product showed a novel previously unreported CBFB-MYH11 fusion (CBFB nt 455-MYH11 nt 1893). Flow cytometric results were analyzed using the Kolmogorov-Smirnov statistic D-value and the median value for positive samples was 0.65 (range, 0.35 to 0.77) versus 0.07 (range, −0.21 to 0.18) in the negative group (P < .0001). The overall concordance between cytogenetics and RT-PCR was 97%, whereas the concordance between flow cytometry and cytogenetics was 100%. Thus, using the anti-inv(16) Ab, all cytogenetically positive and RT-PCR–positive AML cases with inv(16) or t(16;16) could be rapidly identified. This study demonstrates the use of this antibody as an investigational tool in inv(16)/t(16;16) AML and suggests that the development of such reagents may have potential clinical diagnostic use.


2000 ◽  
Vol 74 (3) ◽  
pp. S247
Author(s):  
I Sozen ◽  
L.M Senturk ◽  
L Gutierrez ◽  
E Kovanci ◽  
A Arici

2020 ◽  
Vol 182 (5) ◽  
pp. 1104-1116
Author(s):  
Amanda Barone Pritchard ◽  
Stanley M. Kanai ◽  
Bryan Krock ◽  
Erica Schindewolf ◽  
Jennifer Oliver‐Krasinski ◽  
...  

Zygote ◽  
2019 ◽  
Vol 27 (02) ◽  
pp. 82-88 ◽  
Author(s):  
Vivek Pandey ◽  
Anima Tripathi ◽  
Pawan K. Dubey

SummaryThe decision by germ cells to differentiate and undergo either oogenesis or spermatogenesis takes place during embryonic development and Nanos plays an important role in this process. The present study was designed to investigate the expression patterns in rat of Nanos2-homologue protein in primordial germ cells (PGCs) over different embryonic developmental days as well as in spermatogonial stem cells (SSCs). Embryos from three different embryonic days (E8.5, E10.5, E11.5) and SSCs were isolated and used to detect Nanos2-homologue protein using immunocytochemistry, western blotting, reverse transcription polymerase chain reaction (RT-PCR) and flow cytometry. Interestingly, Nanos2 expression was detected in PGCs at day E11.5 onwards and up to colonization of PGCs in the genital ridge of fetal gonads. No Nanos2 expression was found in PGCs during early embryonic days (E8.5 and 10.5). Furthermore, immunohistochemical and immunofluorescence data revealed that Nanos2 expression was restricted within a subpopulation of undifferentiated spermatogonia (As, single type A SSCs and Apr, paired type A SSCs). The same results were confirmed by our western blot and RT-PCR data, as Nanos2 protein and transcripts were detected only in PGCs from day E11.5 and in undifferentiated spermatogonia (As and Apr). Furthermore, Nanos2-positive cells were also immunodetected and sorted using flow cytometry from the THY1-positive SSCs population, and this strengthened the idea that these cells are stem cells. Our findings suggested that stage-specific expression of Nanos2 occurred on different embryonic developmental days, while during the postnatal period Nanos2 expression is restricted to As and Apr SSCs.


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