• Title, Summary, Keyword: Bal-17

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Differential Functional Expression of Clotrimazole-sensitive $Ca^{2+}$-activated $K^+$ Current in Bal-17 and WEHI-231 Murine B Lymphocytes

  • Zheng, Haifeng;Ko, Jae-Hong;Nam, Joo-Hyun;Earm, Yung-E;Kim, Sung-Joon
    • The Korean Journal of Physiology and Pharmacology
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    • v.10 no.1
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    • pp.19-24
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    • 2006
  • The intermediate conductance $Ca^{2+}-activated$ $K^+$ channels (SK4, IKCa1) are present in lymphocytes, and their membrane expression is upregulated by various immunological stimuli. In this study, the activity of SK4 was compared between Bal-17 and WEHI-231 cell lines which represent mature and immature stages of murine B lymphocytes, respectively. The whole-cell patch clamp with high-$Ca^{2+}$ ($0.8{\mu}M$) KCl pipette solution revealed a voltage-independent $K^+$ current that was blocked by clotrimazole (1 mM), an SK4 blocker. The expression of mRNAs for SK4 was confirmed in both Bal-17 and WEHI-231 cells. The density of clotrimazole-sensitive SK4 current was significantly larger in Bal-17 than WEHI-231 cells ($-11.4{\pm}3.1$ Vs. $-5.7{\pm}1.15$ pA/pF). Also, the chronic stimulation of B cell receptors (BCR) by BCR-ligation (anti-IgM Ab, $3{\mu}g$/ml, 8∼12 h) significantly upregulated the amplitude of clotrimazolesensitive current from $-11.4{\pm}3.1$ to $-53.1{\pm}8.6$ pA/pF in Bal-17 cells. In WEHI-231 cells, the effect of BCR-ligation was significantly small ($-5.7{\pm}1.15$ to $-9.0{\pm}1.00$ pA/pF). The differential expression and regulation by BCR-ligation might reflect functional changes in the maturation of B lymphocytes.

WEHI-231 cells are defective in the ligand-induced internalization of B cell antigen receptor

  • Yoon, Sang Soon;Kim, Tae Jin
    • IMMUNE NETWORK
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    • v.1 no.3
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    • pp.196-202
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    • 2001
  • Backgorund: WEHI-231 B cell line is a representative model for $IgM^+$ mature B cells. To understand the signaling differences between mature and immature B cells, we compared the responsiveness of WEHI-231 and Bal 17 B cell lines to BCR cross-linking. Methods: The extents of tyrosine phosphorylation, ligand-induced internalization, and activation-induced cell death upon BCR cross-linking were compared in two cell lines. Results: Despite a higher expression of BCR, cross-linking of BCR on WEHI-231 cell evoked a weaker level of tyrosine phosphorylation and BCR endocytosis than Bal 17 cells. Furthermore, the endocytosed BCR could not enter the lysosomal compartment and stayed as peripheral spots in WEHI-231 cells. Conclusion: WEHI-231 cell showed preferred BCR-mediated signaling pathways leading to a reduced capability of antigen presentation as well as the enhanced apoptosis in comparision with Bal 17 cells. These results might reflect the signaling differences between mature and immature B cells.

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The Clinical Utility of Polymerase Chain Reaction in the Bronchoalveolar Lavage Fluid for the Detection of Mycobacteria (객담 도말 음성인 환자에서 기관지폐포 세척액 결핵균 중합 효소 연쇄반응 검사의 유용성)

  • Mo, Eun-Kyung;Kyung, Tae-Young;Kim, Dong-Gyu;Park, Myung-Jae;Lee, Myung-Goo;Hyun, In-Gyu;Jung, Ki-Suck;Lee, Kyung-Wha
    • Tuberculosis and Respiratory Diseases
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    • v.45 no.3
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    • pp.519-528
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    • 1998
  • Background: Diagnosis of pulmonary tuberculosis is not easy when the sputum smear for Mycobacterium tuberculosis(M. Tb) is negative. We evaluated the clinical utility of polymerase chain reaction(PCR) for detecting M. Tb in bronchoalveolar lavage(BAL) samples. Methods: We recruited 84 patients whose sputum smear for M. Tb were negative or not available due to no production of sputum. We performed bronchoalveolar lavage for acid-fast stain, culture of mycobacteria, and PCR assay of BAL fluid. We analyzed the results of microbiologic examination. Results: The sensitivity of BAL fluid smear, culture, and PCR were 20%, 38%, and 40%, respectively. The specificity of BAL fluid PCR was 95%. The positive predictive value of PCR was 89%. The smear of BAL fluid was positive in 17%. The PCR of BAL fluid was the only diagnostic test in 17%. Therefore, the BAL fluid analysis including smear and PCR was diagnostic in 34 % within 24 hours. The BAL fluid analysis including smear, PCR, and culture was diagnostic in 55% within 2 month. Conclusion: The BAL fluid PCR was valuable method in the diagnosis of pulmonary tuberculosis in patients whose sputa were not available or reveal negative smear.

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Higher Expression of TRPM7 Channels in Murine Mature B Lymphocytes than Immature Cells

  • Kim, Jin-Kyoung;Ko, Jae-Hong;Nam, Joo-Hyun;Woo, Ji-Eun;Min, Kyeong-Min;Earm, Yung-E;Kim, Sung-Joon
    • The Korean Journal of Physiology and Pharmacology
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    • v.9 no.2
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    • pp.69-75
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    • 2005
  • TRPM7, a cation channel protein permeable to various metal ions such as $Mg^{2+}$, is ubiquitously expressed in variety of cells including lymphocytes. The activity of TRPM7 is tightly regulated by intracellular $Mg^{2+}$, thus named $Mg^{2+}$-inhibited cation (MIC) current, and its expression is known to be critical for the viability and proliferation of B lymphocytes. In this study, the level of MIC current was compared between immature (WEHI-231) and mature (Bal-17) B lymphocytes. In both cell types, an intracellular dialysis with $Mg^{2+}$-free solution (140 mM CsCl) induced an outwardly-rectifying MIC current. The peak amplitude of MIC current and the permeability to divalent cation ($Mn^{2+}$) were several fold higher in Bal-17 than WEHI-231. Also, the level of mRNAs for TRPM7, a molecular correspondence of the MIC channel, was significantly higher in Bal-17 cells. The amplitude of MIC was further increased, and the relation between current and voltage became linear under divalent cation-free conditions, demonstrating typical properties of the TRPM7. The stimulation of B cell receptors (BCR) by ligation with antibodies did not change the amplitude of MIC current. Also, increase of extracellular $[Mg^{2+}]_c$ to enhance the $Mg^{2+}$ influx did not affect the BCR ligation-induced death of WEHI-231 cells. Although the level of TRPM7 was not directly related with the cell death of immature B cells, the remarkable difference of TRPM7 might indicate a fundamental change in the permeability to divalent cations during the development of B cells.

Clinical Utility of Pre-B-Cell Colony-Enhancing Factor in Bronchoalveolar Lavage Fluid of Acute Critical Ill Patients with Lung Infiltrates (폐 침윤을 동반한 급성 중증 환자의 기관지 폐포 세척액에서 측정한 Pre-B-Cell Colony-Enhancing Factor의 임상적 유용성)

  • Lee, Kwangha;Hong, Sang-Bum
    • Tuberculosis and Respiratory Diseases
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    • v.67 no.5
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    • pp.402-408
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    • 2009
  • Background: Pre-B-cell colony enhancing factor (PBEF) has been suggested as a novel biomarker in sepsis and acute lung injury. We measured the PBEF in bronchoalveolar lavage (BAL) fluid of acute critically ill patients with lung infiltrates in order to evaluate the clinical utility of measuring PBEF in BAL fluid. Methods: BAL fluid was collected by bronchoscope from 185 adult patients with lung infiltrates. An enzyme-linked immunosorbent assay was then performed on the collected fluids to measure the PBEF. Results: Mean patient age was 59.9 ${\pm}$14.5 years and 63.8% of patients were males. The mean concentration of PBEF in BAL fluid was 17.5 ${\pm}$88.3 ng/mL, and patients with more than 9 ng/mL of PBEF concentration (n=26, 14.1%) had higher Acute Physiology and Chronic Health Evaluation (APACHE) II and Sequential Organ Failure Assessment (SOFA) scores on the BAL exam day. However, there were no significant differences in clinical characteristics between survivors and non-survivors. In patients with leukocytosis (n=93) seen on the BAL exam day, the linear regression analysis revealed a significant, positive relationship between PBEF and APACHE II ($r^2$=0.06), SOFA score ($r^2$=0.08), Clinical Pulmonary Infection Score ($r^2$=0.05), and plateau pressure in patients on ventilators ($r^2$=0.07) (p<0.05, respectively). In addition, multivariate regression analysis with PBEF as a dependent variable showed that the plateau pressure ($r^2$=0.177, p<0.05) was correlated positively with PBEF. Conclusion: The PBEF level in the BAL fluid may be a useful, new biomarker for predicting the severity of illness and ventilator-induced lung injury in critically ill patients with lung infiltates and leukocytosis.

The Expression of Adhesion Molecules on BAL Cells and Serum Soluble ICAM-1 Level after the Radiotherapy for the Lung Cancer and Its Relationship to the Development of of Radiation Pneumonitis and Fibrosis (방사선 치료후 기관지-폐포세척액내 폐포대식세포 및 임파구의 접착분자발현 변화와 방사선에 의한 폐렴 및 폐섬유증발생의 예측인자로서의 의의)

  • Kim, Dong-Soon;Paik, Sang-Hoon;Choi, Eun-Kyung;Chang, Hye-Sook;Choi, Jung-Eun;Lim, Chae-Man;Koh, Yun-Suck;Lee, Sang-Do;Kim, Woo-Sung;Kim, Won-Dong
    • Tuberculosis and Respiratory Diseases
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    • v.43 no.1
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    • pp.75-87
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    • 1996
  • Background: Lung cancer is the second most frequent malignancy in man in Korea. Surgery is the best treatment modality for non-small cell lung cancer, but most patients were presented in far advanced stage. So radiation therapy(RT) with or without chemotherapy is the next choice and radiation-induced pneumonitis and pulmonary fibrosis is the major limiting factor for the curative RT. Radiation pneumonitis is manifested with fever, cough and dyspnea, 2~3 months after the termination of radiotherpy. Chest X ray shows infiltration, typically limited to the radiation field, but occasionally bilateral infiltration was reported. Also Gibson et al reported that BAL lymphocytosis was found in both lungs, even though the radiation was confined to one lung. The aim of this study is to investigate the change of adhesion molecules expression on BAL cells and serum soluble ICAM-1(sICAM-1) level after the RT and its relationship to the development of radiation pneumonitis. The second aim is to confirm the bilaterality of change of BAL cell pattern and adhesion molecule expression. Subjects: BAL and the measurement of sICAM level in serum and BALF were done on 29 patients with lung cancer who received RT with curative intention. The BAL was done before the RT in 16 patients and 1~2 month after RT in 18 patients. 5 patients performed BAL before and after RT. Result: Clinically significant radiation pneumonitis developed in 7 patients. After RT, total cell count in BAL was significantly increased from $(20.2{\pm}10.2){\times}10^6\;cells/ml$ to $(35.3{\pm}21.6){\times}10^6\;cells/ml$ (p=0.0344) and %lymphocyte was also increased from $5.3{\pm}4.2%$ to $39.6{\pm}23.4%$ (p=0.0001) in all patient group. There was no difference between ipsilateral and contraleteral side to RT, and between the patients with and without radiation-pneumonitis. In whole patient group, the level of sICAM-1 showed no significant change after RT(in serum: $378{\pm}148$, $411{\pm}150\;ng/ml$, BALF: $20.2{\pm}12.2$, $45.1{\pm}34.8\;ng/ml$, respectively), but there was a significant difference between the patients with pneumonitis and without pneumonitis (serum: $505{\pm}164$ vs $345{\pm}102\;ng/ml$, p=0.0253, BALF: $67.9{\pm}36.3$ vs $25.2{\pm}17.9\;ng/ml$, p=0.0112). The expression of ICAM-1 on alveolar macrophages (AM) tends to increase after RT (RMFI: from $1.28{\pm}0.479$ to $1.63{\pm}0.539$, p=0.0605), but it was significantly high in patients with pneumonitis ($2.10{\pm}0.390$) compared to the patients without pneumonitis ($1.28{\pm}0.31$, p=0.0002). ICAM-1 expression on lymphocytes and CD 18 (${\beta}2$-integrin) expression tended to be high in the patients with pneumonitis but the difference was statiastically not significant. Conclusion: Subclinical alveolitis on the basis of BAL finding developed bilaterally in all patients after RT. But clinically significant pneumonitis occurred in much smaller fraction and the ICAM-1 expression on AM and the sICAM-1 level in serum were good indicator of it.

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The Relationship between Cell-mediated Immunity and Subtypes of Lymphocyte in BAL Fluid and Peripheral Blood in Patients with Pulmonary Tuberculosis (폐결핵 환자의 말초 혈액 및 기관지 폐포세척액내의 임파구 아형과 세포성 매개면역과의 관계)

  • Uh, Soo-Taek;Cha, Mi-Kyung;Lee, Sang-Moo;Kim, Hyun-Tae;Chung, Yeon-Tae;Woo, Jun-Hee;Kim, Yong-Hun;Park, Choon-Sik
    • Tuberculosis and Respiratory Diseases
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    • v.39 no.4
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    • pp.334-342
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    • 1992
  • Background: The activated T lymphocyte by inhalaed mycobacterial antigen may evoke cell-mediated immunity in patients with active pulmonary tuberculosis. These activated lymphocyte may influence the response of tuberculin-purified protein derivative (PPD) in skin test. But occasionally, anergy to PPD appear in patients with pulmonary tuberculosis in spite of active stage. Thus we evaluated the effect of change of subtypes of lymphocyte in bronchoalveolar lavage fluid (BAL) and peripheral blood on anergy to PPD in patients with active pulmonary tuberculosis. Method: We performed tuberculin skin test and flow-cytometry analysis of lymphocytes obtained from BAL fluid and peripheral blood in 11 healthy normal volunteers and 20 patients with active pulmonary tuberculosis. Results: 1) The composition of lymphocyte significantly increased in patients with active pulmonary tuberculosis when compared with that in healthy control ($25.2{\pm}4.8$ vs $6.5{\pm}1.3%$, p<0.01), but composition of monocyte significantly decreased ($69.6{\pm}5.7$ vs $89.2{\pm}1.4%$, p<0.05) in analysis of BAL fluid. 2) There were no differences in compositions of cells in BAL fluid between responders and no-responders to PPD. 3) The compositions of CD3 (+), CD4 (+), CD3 (+) IL-2R (+), CD3 (+) HLA-DR (+) significantly increased in BAL fluid when compared with those in peripheral blood in patients with active pulmonary tuberculosis. But the composition of CDS (+), CD4/CDS were not different between BAL fluid and peripheral blood. 4) There were no correlations between response to PPD and compositions of cells and lymphocyte subtypes in BAL fluid and peripheral blood in all patients with tuberculosis, responders, and no-responders, respectively. Conclusion: From these results, we suggest no direct relationship between compositions of inflammatory cells in bronchoalveolar lavage fluid and we could not rule out the possibility of compartmentalization of activated lymphocyte involving in anergy to PPD in skin test in patients with active pulmonary tuberculosis.

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The Role of Protein Kinase C in Acute Lung Injury Induced by Endotoxin (내독소에 의한 급성폐손상에서 Protein Kinase C의 역할)

  • Kim, Yong-Hoon;Moon, Seung-Hyug;Kee, Sin-Young;Ju, Jae-Hak;Park, Tae-Eung;Im, Keon-Il;Cheong, Seung-Whan;Kim, Hyeon-Tae;Park, Choon-Sik;Jin, Byung-Won
    • Tuberculosis and Respiratory Diseases
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    • v.44 no.2
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    • pp.349-359
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    • 1997
  • Background : The signal pathways and their precise roles for acute respiratory distress syndrome caused by endotoxin (ETX) has not been established. Since there has been several in vitro experiments suggesting that activation of protein kinase C (PKC) pathway may be responsible for endotoxin-induced inflammatory reaction, we performed in vivo experiments in the rats with the hypothesis that PKC-inhibition can effectively prevent endotoxin-induced acute lung injury. Methods : We studied the role of PKC in ETX-induced ALI using PKC inhibitor (staurosporine, STP) in the rat Specific pathogen free male Sprague-Dawley weighted from 165 to 270g were used for the study. Animals were divided into the normal control (NC)-, vehicle control (VC)-, ETX-, PMA (phorbolmyristateacetate)-, STP+PMA-, and STP+ETX-group. PMA (50mg/kg) or ETX (7mg/kg) was instilled through polyethylen catheter after aseptic tracheostomy with and without STP (0.2mg/kg)-pretreatment STP was injected via tail vein 30min before intratracheal injection (IT) of PMA or ETX. Bronchoalveolar lavage (BAL) was done 3-or 6-hrs after IT of PMA or ETX respectively, to measure protein concentration, total and differential cell counts. Results : The results were as follows. The protein concentrations in BALF in the PMA- and ETX-group were very higher than that of VC-group (p<0.001). When animals were pretreated with STP, the %reduction of the protein concentration in BALF was $64.8{\pm}8.5$ and $30.4{\pm}2.5%$ in the STP+PMA- and STP+ETX-group, respectively (p = 0.028). There was no difference in the total cell counts between the PMA-and VC-group (p = 0.26). However the ETX-group showed markedly increased total cell counts as compared to the VC- (p = 0.003) and PMA-group (p = 0.0027), respectively. The total cell counts in BALF were not changed after pretreatment with STP compared to the PMA- (p = 0.22) and ETX-group (p = 0.46). The percentage of PMN, but not alveolar macrophage, was significantly elevated in the PMA-, and ETX-group. Especially in the ETX-group, the percentage of PMN was 17 times higher than that of PMA (p < 0.001). The differential cell counts was not different between the PMA and STP+PMA On the contrary the STP+ETX-group showed decreased percentage of PMN (p = 0.016). There was no significant relationship between the protein concentration and the total or differential cell counts in each group. Conclusion : Pretreatment with PKC-inhibitor (staurosporine) partially but significantly inhibited ETX-induced ALI.

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CD40 Co-stimulation Inhibits Sustained BCR-induced $Ca^{2+}$ Signaling in Response to Long-term Antigenic Stimulation of Immature B Cells

  • Nguyen, Yen Hoang;Lee, Ki-Young;Kim, Tae-Jin;Kim, Sung-Joon;Kang, Tong-Mook
    • The Korean Journal of Physiology and Pharmacology
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    • v.15 no.3
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    • pp.179-187
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    • 2011
  • Regulation of B cell receptor (BCR)-induced $Ca^{2+}$ signaling by CD40 co-stimulation was compared in long-term BCR-stimulated immature (WEHI-231) and mature (Bal-17) B cells. In response to long-term pre-stimulation of immature WEHI-231 cells to ${\alpha}$-IgM antibody (0.5~48 hr), the initial transient decrease in BCR-induced $[Ca^{2+}]_i$ was followed by spontaneous recovery to control level within 24 hr. The recovery of $Ca^{2+}$ signaling in WEHI-231 cells was not due to restoration of internalized receptor but instead to an increase in the levels of $PLC{\gamma}2$ and $IP_3R-3$. CD40 co-stimulation of WEHI-231 cells prevented BCR-induced cell cycle arrest and apoptosis, and it strongly inhibited the recovery of BCR-induced $Ca^{2+}$ signaling. CD40 co-stimulation also enhanced BCR internalization and reduced expression of $PLC{\gamma}2$ and $IP_3R-3$. Pre-treatment of WEHI-231 cells with the antioxidant N-acetyl-L-cysteine (NAC) strongly inhibited CD40-mediated prevention of the recovery of $Ca^{2+}$ signaling. In contrast to immature WEHI-231 cells, identical long-term ${\alpha}$-IgM pre-stimulation of mature Bal-17 cells abolished the increase in BCR-induced $[Ca^{2+}]_i$, regardless of CD40 co-stimulation. These results suggest that CD40-mediated signaling prevents antigen-induced cell cycle arrest and apoptosis of immature B cells through inhibition of sustained BCR-induced $Ca^{2+}$ signaling.

Development of molecular markers among Barred Plymouth rock, Korean Ogol Chicken and White Leghorn

  • Choi, Jin-Won;Lee, Eun-Young;Lee, Jae-Hee;Kim, Duk-Kyung;Kim, Hee-Bal;Han, Jae-Yong
    • Proceedings of the Korea Society of Poultry Science Conference
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    • pp.68-69
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    • 2005
  • To identify germline chimeric chicken using germ cell transplantation method, the testcross, spends much time, labor and cost to perform, is the only way for distinguishing germline chimeric chicken from normal one And to enhance the method, development of breed-specific molecular markers have been needed. We have just identified breed-specific sequence polymorphisms among Barred Plymouth rock, Korean Ogol Chicken and White Leghorn in PMEL17 and MC1R gene the loci of which are identical to dominant white and extended black loci. These sequence polymorphism will be very useful for screening germline chimera.

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