• Title/Summary/Keyword: reverse transcriptase polymerase chain reaction

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Comparison for immunophysiological responses of Jeju and Thoroughbred horses after exercise

  • Khummuang, Saichit;Lee, Hyo Gun;Joo, Sang Seok;Park, Jeong-Woong;Choi, Jae-Young;Oh, Jin Hyeog;Kim, Kyoung Hwan;Youn, Hyun-Hee;Kim, Myunghoo;Cho, Byung-Wook
    • Asian-Australasian Journal of Animal Sciences
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    • 제33권3호
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    • pp.424-435
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    • 2020
  • Objective: The study was conducted to investigate variations in the immunophysiological responses to exercise-induced stress in Jeju and Thoroughbred horses. Methods: Blood samples were collected from the jugular veins of adult Jeju (n = 5) and Thoroughbred (n = 5) horses before and after 30 min of exercise. The hematological, biochemical, and immunological profiles of the blood samples were analyzed. Blood smears were stained and observed under a microscope. The concentration of cell-free (cf) DNA in the plasma was determined using real time polymerase chain reaction (PCR). Peripheral blood mononuclear cells (PBMCs) and polymorphonuclear cells were separated using Polymorphprep, and the expression of various stress-related and chemokine receptor genes was measured using reverse transcriptase (RT) and real-time PCR. Results: After exercise, Jeju and Thoroughbred horses displayed stress responses with significantly increased rectal temperatures, cortisol levels, and muscle catabolism-associated metabolites. Red blood cell indices were significantly higher in Thoroughbred horses than in Jeju horses after exercise. In addition, exercise-induced stress triggered the formation of neutrophil extracellular traps (NETs) and reduced platelet counts in Jeju horses but not in Thoroughbred horses. Heat shock protein 72 and heat shock protein family A (Hsp70) member 6 expression is rapidly modulated in response to exercise-induced stress in the PBMCs of Jeju horses. The expression of CXC chemokine receptor 4 in PBMCs was higher in Thoroughbred horses than in Jeju horses after exercise. Conclusion: In summary, the different immunophysiological responses of Jeju and Thoroughbred horses explain the differences in the physiological and anatomical properties of the two breeds. The physiology of Thoroughbred horses makes them suitable for racing as they are less sensitive to exercise-induced stress compared to that of Jeju horses. This study provides a basis for investigating the link between exercise-induced stresses and the physiological alteration of horses. Hence, our findings show that some of assessed parameters could be used to determine the endurance performance of horses.

Characterization of Acetylcholine-induced Currents in Male Rat Pelvic Ganglion Neurons

  • Park, Joong-Hyun;Park, Kyu-Sang;Cha, Seung-Kyu;Lee, Keon-Il;Kim, Min-Jung;Park, Jong-Yeon;Kong, In-Deok;Lee, Joong-Woo
    • The Korean Journal of Physiology and Pharmacology
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    • 제8권4호
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    • pp.219-225
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    • 2004
  • The pelvic ganglia provide autonomic innervations to the various urogenital organs, such as the urinary bladder, prostate, and penis. It is well established that both sympathetic and parasympathetic synaptic transmissions in autonomic ganglia are mediated mainly by acetylcholine (ACh). Until now, however, the properties of ACh-induced currents and its receptors in pelvic ganglia have not clearly been elucidated. In the present study, biophysical characteristics and molecular nature of nicotinic acetylcholine receptors (nAChRs) were studied in sympathetic and parasympathetic major pelvic ganglion (MPG) neurons. MPG neurons isolated from male rat were enzymatically dissociated, and ionic currents were recorded by using the whole cell variant patch clamp technique. Total RNA from MPG neuron was prepared, and RT-PCR analysis was performed with specific primers for subunits of nAChRs. ACh dose-dependently elicited fast inward currents in both sympathetic and parasympathetic MPG neurons $(EC_{50};\;41.4\;{\mu}M\;and\;64.0\;{\mu}M,\;respectively)$. ACh-induced currents showed a strong inward rectification with a reversal potential near 0 mV in current-voltage relationship. Pharmacologically, mecamylamine as a selective antagonist for ${\alpha}3{\beta}4$ nAChR potently inhibited the ACh-induced currents in sympathetic and parasympathetic neurons $(IC_{50};\;0.53\;{\mu}M\;and\;0.22\;{\mu}M,\;respectively)$. Conversely, ${\alpha}-bungarotoxin$, ${\alpha}-methyllycaconitine$, and $dihydro-{\beta}-erythroidine$, which are known as potent and sensitive blockers for ${\alpha}7$ or ${\alpha}4{\beta}2$ nAChRs, below micromolar concentrations showed negligible effect. RT-PCR analysis revealed that ${\alpha}3$ and ${\beta}4$ subunits were predominantly expressed in MPG neurons. We suggest that MPG neurons have nAChRs containing ${\alpha}3$ and ${\beta}4$ subunits, and that their activation induces fast inward currents, possibly mediating the excitatory synaptic transmission in pelvic autonomic ganglia.

Stimulative Effects of Hominis Placental Pharmacopuncture Solution Combined with Zinc-oxide Nanoparticles on RAW 264.7 Cells - ZnO HPPS more easily stimulates RAW 264.7 cells -

  • Hong, Tae-Keun;Kim, Jee-Hye;Woo, Ju-Youn;Ha, Ki-Tae;Joo, Myung-Soo;Hahn, Yoon-Bong;Jeong, Han-Sol
    • 대한약침학회지
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    • 제15권3호
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    • pp.13-18
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    • 2012
  • Objectives: The purpose of this study is to examine whether Hominis Placental pharmacopuncture solution (HPPS) combined with zinc-oxide nanoparticles (ZnO NP) activates RAW 264.7 cells. Methods: We soaked ZnO nanoparticles in the Hominis Placenta pharmacopuncture solution, thereby making a combined form (ZnO NP HPPS). The effect of ZnO NP HPPS on the intracellular reactive oxygen species (ROS) production was measured by 2', 7'-dichlorofluorescin diacetate (DCFH-DA) assay. The effect of ZnO NP HPPS on NF-${\kappa}B$ was measured by using a luciferase assay. The effect of ZnO NP HPPS on the cytokine expression was assessed by semi-quantitative reverse transcriptase polymerase chain reaction (RT-PCR). The cellular uptake of ZnO NP HPPS was measured by using a flow cytometric analysis, and cellular structural alterations were analyzed by using transmission electron microscopy (TEM). Results: Neither the HPPS nor the ZnO NPs induced intracellular ROS production in RAW 264.7 cells. Neither of the materials activated NF-${\kappa}B$ or it's dependent genes, such as TNF-${\alpha}$, IL-1, and MCP-1. However, ZnO NP HPPS, the combined form of ZnO NPs and HPPS, did induce the intracellular ROS production, as well as prominently activating NF-${\kappa}B$ and it's dependent genes. Also, compared to ZnO NPs, it effectively increa-sed the uptake by RAW 264.7 cells. In addition, cellular structural alterations were observed in groups treated with ZnO NP HPPS. Conclusions: Neither ZnO NP nor HPPS activated RAW 264.7 cells, which is likely due to a low cellular uptake. The ZnO NP HPPS, however, significantly activated NF-${\kappa}B$ and up-regulated its dependent genes such as TNF-${\alpha}$, IL-1, and MCP-1. ZnO NP HPPS was also more easily taken into the RAW 264.7 cells than either ZnO NP or HPPS.

신수혈의 침자극과 황기약침이 실험용 생쥐의 면역활성물질인 cytokine의 IL-6 발현에 미치는 영향 (Effect of Acupuncture and Radix Astragali aqua-acupuncture at Synsu(BL23) on transcriptional expression of mouse cytokine IL-6)

  • 김종수;신상습;김철호;박선동;박원환
    • Journal of Acupuncture Research
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    • 제15권2호
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    • pp.147-155
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    • 1998
  • Acupuncture and Radix Astragali aqua-acupuncture stimuli have long been used to cure human diseases. However, it still remains to be unkown on its action mechanism, physiolosical and biochemical aspects. Thus, many attempts were made to show the scientific background covering the above mentioned mechanisms. Most recent studies show that these tests improve blood circulatory system and increase leucocyte counts. In this study, we have applied the acupuncture stimuli to mouse Sinsu(BL-23), which is a stimulative point of oriental medicine, to see if cytokine such as IL-6 can be detected. Mice were treated with lipopolysaccharide(LPS) for inflammation induction, and then reverse transcriptase-polymerase chain reaction (RT-PCR) using each primer set was performed to trace the amounts of mRNA. The results are summarized as follows ; 1. IL-6 was not temporarily expressed in normal mice 15 min after the acupuncture was pulled out. But, it started to show a feeble expression at 30 min after the removal of acupuncture and it started to reduce at 1h. after the acupuncture was pulled out 2. IL-6 was specifically expressed in LPS-treated mouse 30 min after the acupuncture was pulled out. The transcriptional expressions of LPS-treated mice were more effective than those of normal mice at 30 min after the removal of acupuncture 3. IL-6 was not temporarily expressed in normal mice 15 min after Radix Astragali aqua-acupuncture. But it expressed most highly at 30 min, and the transcriptional expressions of IL-6 was continued to 3 h. 4. IL-6 was not expressed in all the time after Radix Astragali aqua-acupuncture in LPS-treated mice. Therefore, a follow-up of cytokine IL-6 can be used not only a basis of the effect of acupuncture and Radix Astragali aqua-acupuncture but a diagnosis giude through the immunological action of thats. And, it is suggested that cytokine's expression by Acupuncture and Radix Astragali aqua-acupuncture stimulation should be continuously elucidated.

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접골목(接骨木)추출물 및 약침액에 의한 MIF 활성 조절능이 생쥐의 제2형 Collagen 유발 관절염에 미치는 영향 (Effect of Suppressing the Activation of Macrophage Migration Inhibitory Factor by $Sambucus$ $williamsii$ $H_{ANCE}$ Extract & Pharmacopuncture Solution on Type II Collagen-induced Arthritis)

  • 이동건;김은정;이은솔;왕개하;조현석;이승덕;김갑성;김경호
    • Journal of Acupuncture Research
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    • 제29권1호
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    • pp.103-113
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    • 2012
  • Objectives : The purpose of this study is to evaluate effect of suppressing the expression of cyclo-oxygenase-type-2 (COX-2) as a consequence of inhibition macrophage migration inhibitory factor (MIF) activation by $Sambucus$ $williamsii$ $Hance$ (SWH) pharmacopunctureon rheumatoid arthritis (RA). Methods : In vitro test, synoviocytes extracted from type II collagen-induced arthritis (CIA) mouse's knee joint were cultivated After that, each well of synoviocytes was mixed with the extract of SWH at the dosage of $0.4mg/m{\ell}$, $0.6mg/m{\ell}$, $0.8mg/m{\ell}$, and $1.0mg/m{\ell}$ respectively, and cultivated for 24 hours after the addition. Reverse transcriptase - polymerase chain reaction (RT-PCR) is used to investigate the expression of MIF, Tumor necrosis factor (TNF)-${\alpha}$, COX-2 mRNA. $In$ $vivo$ test, thirty DBA female mice were used, and each ten mice were allocated into three group; normal group, CIA-elicitated group, and group treated with SWH pharmacopuncture on it's the point of $ST_{35}$ after CIA elicitation. It is investigated that change of mice foot thickness, histologic change of sliced synovial joint of mouse, and extent change of MIF, TNF-${\alpha}$, COX-2 in synovial membrane. Results : $In$ $vitro$ test, the expressions of cytokine(MIF, TNF-${\alpha}$, COX-2) mRNAs related to RA were dose-dependent decreased. In the SWH pharmacopuncture group, foot thickness and histologic change of sliced synovial joint were decreased comparing with CIA-elicitated group's change. In the SWH pharmacopuncture group, the suppression of MIF, TNF-${\alpha}$, COX-2 in synovial membrane was clearly shown comparing with CIA-elicitated group's change. Conclusions : It might be suggested that SWH pharmacopuncture mitigate tissue damage originated from rheumatoid arthritis by suppressing the expression of COX-2 as a consequence of inhibition MIF activation.

신종 인플루엔자 A (H1N1) 감염으로 입원한 성인 폐렴 환자의 임상양상 (Clinical Features of Hospitalized Adult Patients with Pneumonia in Novel Influenza A (H1N1) Infection)

  • 한창훈;현여경;최유리;성나영;박윤선;이꽃실;정재호
    • Tuberculosis and Respiratory Diseases
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    • 제69권1호
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    • pp.24-30
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    • 2010
  • Background: A novel 2009 influenza A (H1N1) virus emerged and disseminated to all over the world. There are few reports on the clinical characteristics of patients with complications. We describe the clinical features of pneumonia in adult patients hospitalized, who have novel influenza infection. Methods: There were 43 adult patients enrolled into the study with pneumonia of 528 hospitalized patients confirmed influenza A (H1N1) virus infection by real-time reverse transcriptase polymerase chain reaction testing, between 24 August 2009 and 31 January 2010. The clinical data of patients with pneumonia were collected retrospectively. Results: There were 22 of 43 (51.2%) influenza patients with pneumonia that had higher risk factors for complications. Compared to 28 patients with influenza A (H1N1) viral pneumonia and 15 patients, who had isolated bacteria from cultures, those with mixed viral and bacterial pneumonia were significantly more likely to have unilobar consolidations on chest radiographs (53.3 vs. 10.7%, p<0.01) and higher scores of pneumonia severity index (PSI; 90 [66~100] vs. 53 [28~90], p=0.04). Six patients required mechanical ventilation support in an Intensive Care Unit and were more likely to have dyspnea (83.3 vs. 29.3%, p=0.02) and low levels of $PaO_2$ (48.3 [37.0~70.5] vs 64.0 [60.0~74.5] mm Hg, p=0.02) and high levels of pneumonia severity index (PSI) score (108.0 [74.5~142.8] vs. 56.0 [40.5~91.0], p=0.03). Conclusion: The majority of pneumonia patients infected with novel influenza improved. Chest radiographic findings of unilobar consolidations suggest that mixed pneumonia is more likely. Initial dyspnea, hypoxemia, and high levels of PSI score are associated with undergoing mechanical ventilation support.

Expression Levels of Vascular Endothelial Growth Factors A and C in Patients with Peptic Ulcers and Gastric Cancer

  • Taghizadeh, Shirin;Sankian, Mojtaba;Ajami, Abolghasem;Tehrani, Mohsen;Hafezi, Nasim;Mohammadian, Rajeeh;Farazmandfar, Touraj;Hosseini, Vahid;Abbasi, Ali;Ajami, Maryam
    • Journal of Gastric Cancer
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    • 제14권3호
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    • pp.196-203
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    • 2014
  • Purpose: Vascular endothelial growth factor (VEGF) is one of the most important growth factors for metastatic tumors. To clarify the role of VEGF-A and C in patients with peptic ulcer disease (PUD) or gastric cancer (GC), we evaluated the expression levels of these two molecules. We also analyzed the effect of Helicobacter pylori infection on VEGF-A and C expression levels. Materials and Methods: Patients with dyspepsia who needed diagnostic endoscopy were selected and divided into three groups: nonulcer dyspepsia (NUD), PUD, and GC, according to their endoscopic and histopathological results. Fifty-two patients with NUD, 50 with PUD, and 38 with GC were enrolled in this study. H. pylori infection was diagnosed by the rapid urease test. After RNA extraction and synthesis of cDNA, the expression levels of VEGF-A and C were determined by quantitative reverse transcriptase polymerase chain reaction. Results: The VEGF-C expression level in the PUD and GC groups was significantly higher than that in the NUD group. Moreover, the VEGF-A expression level in the PUD and GC groups was higher than in the NUD group, although the differences were not statistically significant. Significant positive correlations were also observed between the expression levels of these two molecules in the PUD and GC groups. In addition, the expression levels of these two molecules were higher in H. pylori positive patients with PUD or GC than in H. pylori negative patients of the same groups; however, these differences did not reach statistical significance. Conclusions: Up-regulation of VEGF-C expression during gastric mucosal inflammation may play a role in the development of peptic ulcers or GC.

The Inhibitory Effect of Chlorophyllin is Influenced by Different Promotion Stages in DMBA-TPA-induced Mouse Skin Carcinogenesis

  • Kim, Jin;Yook, Jong-In;Park, Kwang-Kyun;Lee, Eun-Ha;Jung, So-Young;Joon, Yin-Liu;Kyung, Chul-Hong;Kim, Ju;Chung, Won-Yoon
    • 한국환경성돌연변이발암원학회지
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    • 제19권1호
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    • pp.46-55
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    • 1999
  • To develop a chemopreventive strategy based on the different stages of premalignant lesions, we hypothesized that the inhibitory effect of chemopreventive agents is influenced by different promotion stages during carcinogenesis. DMBA-TPA-induced skin carcinogenesis was used with ICR mice and chlorophyllin (CHL) was applied as a chemopreventive agent. In vitro assay was performed with Salmonella typhi. TA100 to observe any anti-mutagenic activity of CHL against DMBA. Pre-initiation and pre-promotion effects of CHL were observed by CHL treatment before initiation and before promotion. To evaluate the inhibitory effect at different promotion stages, each group was divided into three subgroups after TPA promotion for 6, 18 and 24 weeks, respectively ; the first subgroup was immediately sacrificed after termination of TPA, the second subgroup was treated with CHL, and the third subgroup was sacrificed 8 weeks after termination of TPA without CHL treatment. The degrees of epithelial dysplasia, papilloma formation, and invasive carcinoma were observed histologically, and GST-Pi expression was observed immunohistochemically. ODC mRNA level was analyzed by reverse transcriptase-polymerase chain reaction. Results showed : CHL dose-dependently inhibited the mutation of Salmonella typhi. TA100; the incidence of epithelial dysplasia and papilloma formation was lower in pre-initiation and pre-promotion CHL-treated mice than DMBA-TPA-treated mice; no invasive carcinoma developed in pre-initiation CHL-treated groups, while 67% of DMBA-TPA treated mice had carcinomas; GST-Pi expression decreased when CHL was treated before initiation and before promotion; and when CHL was treated after termination of TPA application at 18 and 24- week-TPA promotion stages, respectively, the incidence of epithelial dysplasia and papilloma was markedly reduced compared to non-treated groups. When comparing the incidence of epithelial dysplasia and papilloma between the immediately-sacrificed subgroup and the non-treated group with a waiting period, we speculated that the 18-week-TPA promotion stage might belong to the promoter-independent progression stage. At the 18-week-TPA promotion stage, the level of ODC mRNA in the CHL-treated group was clearly reduced to the level of normal tissue. Taking these results together, CHL showed both anti-initiation and anti-promotion effects, while the inhibitory effect of CHL was prominent in the 18-week-TPA promotion stage. However, CHL seems to be incapable of completely blocking the progression in the 24-week-TPA promotion stage.

Clinical characteristics and outcomes among pediatric patients hospitalized with pandemic influenza A/H1N1 2009 infection

  • Lee, Eun;Seo, Ju-Hee;Kim, Hyung-Young;Na, Shin;Kim, Sung-Han;Kwon, Ji-Won;Kim, Byoung-Ju;Hong, Soo-Jong
    • Clinical and Experimental Pediatrics
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    • 제54권8호
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    • pp.329-334
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    • 2011
  • Purpose: The purpose of this article is to describe the clinical and epidemiologic features and outcomes among children hospitalized with pandemic influenza A/H1N1 2009 infection. Methods: We retrospectively reviewed the charts of hospitalized pediatric patients (<18 years) diagnosed with pandemic influenza A/H1N1 2009 infection by reverse-transcriptase polymerase chain reaction at a tertiary hospital in Seoul, Korea, between September 2009 and February 2010. Results: A total of 72 children were hospitalized with pandemic influenza A/H1N1 2009 infection (median age, 6.0 years; range, 2 months to 18 years). A total of 40% had at least 1 underlying medical condition, including asthma (17%), malignancies (19%), and heart diseases (17%). Of the 72 patients, 54 (76%) children admitted with H1N1 infection showed radiographic alterations compatible with pneumonia. There was no significant difference in pre-existing conditions between pandemic influenza A/H1N1 infected patients with or without pneumonia. Children with pandemic influenza A/H1N1 pneumonia were more likely to have a lower lymphocyte ratio (P=0.02), higher platelet count (P=0.02), and higher level of serum glucose (P=0.003), and more commonly presented with dyspnea than did those without pneumonia (P=0.04). Conclusions: No significant differences in age, sex, or presence of preexisting conditions were found between children hospitalized with pandemic influenza A/H1N1 H1N1 influenza infection with pneumonia and those without pneumonia. Higher leukocyte count, higher glucose level, and a lower lymphocyte ratio were associated with the development of pandemic A/H1N1 2009 influenza pneumonia.

소아에서 human metapneumovirus 감염의 임상 특징 (Clinical Manifestation of Human Metapneumovirus Infection in Korean Children)

  • 안정민;최성열;김동수;김기환
    • Pediatric Infection and Vaccine
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    • 제20권1호
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    • pp.28-35
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    • 2013
  • 목 적 : 2000년이후human metapneumovirus (hMPV)는 하기도 호흡기 감염을 일으키는 주요 바이러스로 인식되고 있다. 급성 호흡기 감염으로 입원한 소아 환자들에서의 hMPV의 역학 및 임상적 특징, 그리고 혈액학적 특징 및 hMPV에 감염되지 않은 환자와의 비교, 중복 감염등을 통해 hMPV의 특징을 파악하고자 하였다. 방 법 : 2010년 2월부터 2012년 1월까지 급성 호흡기 증상으로 입원한 환아 1,554명의 비인두 흡인을 통해 객담 검체를 채취하여 RT-PCR을 시행 후 hMPV가 검출된 99명의 환아들의 임상 기록지를 후향적으로 검토하여 이들의 임상 증상 및 검사 소견을 분석 비교 하였다. 결 과 : 1,554명 중 1,004명(64.6%)에서 호흡기 바이러스가 검출되었다. 그 중 hMPV가 양성인 환자는 99명으로 양성 검체의 9.8%였다. hMPV에 감염된 환자의 평균 연령은 25.0개월이었으며 해마다 유행 정도는 다르나 4월부터 6월에 주로 발현하였으며 발열기간이 다른 바이러스 보다 길고 호흡기 증중을 보였으며, 천명(wheezing)이 특징적 임상 증상이었다. 진단으로 폐렴이 가장 많았지만, 12개월 이하의 연령군에서는 모세기관지염이 가장 흔하였다. 혈액학적 검사소견은 ESR (erythrocyte sedimentation rate), CRP (C-reactive protein) 모두 hMPV에 감염된 환자군에서 정상 범위 보다 상승된 소견을 보였다. hMPV의 단독 감염과 중복 감염을 비교해 보았을 때 임상증상 및 혈액 검사 소견에서 통계학적 차이를 보이지 않았다. 결론 : hMPV는 폐렴을 일으키는 주요 원인 바이러스이며 13개월 이상부터 48개월 이하의 연령에서 호발하며 평균 연령은 25개월이며 4월에서 6월까지 유행하는 양상을 보였다. 천명이 임상적 특징이며, hMPV에 대하여 우리나라에서의 보고는 점차 증가되고 있으나 아직은 생물학적, 유전학적 특징과 관련된 임상 분석이 더 필요하다.

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