scholarly journals Molecular Cloning, Expression, and Immunogenicity of MTB12, a Novel Low-Molecular-Weight Antigen Secreted byMycobacterium tuberculosis

1998 ◽  
Vol 66 (9) ◽  
pp. 4208-4214 ◽  
Author(s):  
John R. Webb ◽  
Thomas S. Vedvick ◽  
Mark R. Alderson ◽  
Jeffrey A. Guderian ◽  
Shyian S. Jen ◽  
...  
Keyword(s):  
Molecular Weight ◽  
Molecular Cloning ◽  
Mononuclear Cells ◽  
Single Copy ◽  
Low Molecular Weight ◽  
Peripheral Blood Mononuclear ◽  
Purified Protein Derivative ◽  
Culture Filtrate Proteins ◽  
Processed Form

ABSTRACT Proteins secreted into the culture medium by Mycobacterium tuberculosis are thought to play an important role in the development of protective immune responses. In this report, we describe the molecular cloning of a novel, low-molecular-weight antigen (MTB12) secreted by M. tuberculosis. Sequence analysis of the MTB12 gene indicates that the protein is initially synthesized as a 16.6-kDa precursor protein containing a 48-amino-acid hydrophobic leader sequence. The mature, fully processed form of MTB12 protein found in culture filtrates has a molecular mass of 12.5 kDa. MTB12 protein constitutes a major component of the M. tuberculosis culture supernatant and appears to be at least as abundant as several other well-characterized culture filtrate proteins, including members of the 85B complex. MTB12 is encoded by a single-copy gene which is present in both virulent and avirulent strains of the M. tuberculosis complex, the BCG strain of M. bovis, and M. leprae. Recombinant MTB12 containing an N-terminal six-histidine tag was expressed in Escherichia coli and purified by affinity chromatography. Recombinant MTB12 protein elicited in vitro proliferative responses from the peripheral blood mononuclear cells of a number of purified protein derivative-positive (PPD+) human donors but not from PPD− donors.

scholarly journals Coordinated In Vitro Release of Granulysin, Perforin and IFN-γ in TB and HIV/TB Co-Infection Associated with Clinical Outcomes before and after Anti-TB Treatment

Pathogens ◽  
2020 ◽  
Vol 9 (8) ◽  
pp. 655
Author(s):  
Nada Pitabut ◽  
Panadda Dhepakson ◽  
Shinsaku Sakurada ◽  
Naoto Keicho ◽  
Srisin Khusmith
Keyword(s):  
Clinical Outcomes ◽  
Mononuclear Cells ◽  
Granzyme B ◽  
In Vitro Release ◽  
Peripheral Blood Mononuclear ◽  
Purified Protein Derivative ◽  
Tb Treatment ◽  
Before And After ◽  
Ifn Γ

Granule-associated killing molecules released from cytotoxic T lymphocytes participate as a crucial step in immunity against tuberculosis (TB), but the role of coordinated production remains controversial. Coordinated release of effector molecules in vitro after stimulating peripheral blood mononuclear cells (PBMCs) of active TB or HIV/TB coinfection patients with PPD, purified protein derivative of tuberculin and avirulent Mtb, H37Ra, an attenuated strain were investigated in association with clinical outcomes. Perforin, granzyme-B, granulysin and IFN-γ were measured using ELISA. Before anti-TB treatment, PBMCs of TB stimulated with PPD or H37Ra released higher perforin, granzyme-B, and granulysin levels than in HIV/TB and released significantly higher IFN-γ (p = 0.045, p = 0.022). Granulysin positively correlated with perforin in TB (p = 0.042, r = 0.385), HIV/TB coinfection (p = 0.003, r = 0.941) after PPD stimulation, and after H37Ra stimulation in TB (p = 0.005, r = 0.549), but negatively correlated with granzyme B in TB (p = 0.042, r = −0.386), HIV/TB coinfection (p = 0.042, r = 0.754) were noted. After anti-TB treatment, increased levels of perforin, granulysin and IFN-γ in TB or HIV/TB upon PPD or H37Ra stimulation, and decreased granzyme-B levels after PPD (p = 0.003) or H37Ra (p = 0.028) stimulation in TB were observed. These results suggest that granulysin may act synergistic with perforin and IFN-γ in TB, indicating its crucial function in host immunity to tuberculosis. Future studies with larger numbers of patients ought to be conducted in the future.

Fibrinogen and fibrin induce synthesis of proinflammatory cytokines from isolated peripheral blood mononuclear cells

2007 ◽  
Vol 97 (05) ◽  
pp. 822-829 ◽  
Author(s):  
Peter Kierulf ◽  
Per Sandset ◽  
Olav Klingenberg ◽  
Gunn Joø ◽  
Hans Godal ◽  
...  
Keyword(s):  
Molecular Weight ◽  
Acute Phase ◽  
Proinflammatory Cytokines ◽  
Peripheral Blood Mononuclear Cells ◽  
Peripheral Blood ◽  
Mononuclear Cells ◽  
Low Molecular Weight ◽  
Peripheral Blood Mononuclear ◽  
Tnf Α ◽  
Blood Mononuclear Cells

SummaryFibrinogen in plasma includes three main fractions; high-molecular- weight (HMW) -fibrinogen, low-molecular-weight (LMW) -fibrinogen, and very-low-molecular-weight (LMW`) -fibrinogen. During acute-phase conditions, plasma fibrinogen levels and the HMW-/LMW-fibrinogen ratio increase rapidly due to increased synthesis of HMW-fibrinogen. The consequences of elevated plasma fibrinogen levels and local deposition of fibrin in inflammatory tissues observed during acute-phase conditions are not clear.We wanted to investigate proinflammatory effects of fibrinogen and fibrin on peripheral blood mononuclear cells (PBMC) as reflected by altered mRNA expression and synthesis of the proinflammatory cytokines IL-6,TNF- α and IL-1 β, and to explore the significance of altered HMW-/LMW-fibrinogen ratio. PBMC were isolated from whole blood using Lymphoprep® . HMW-fibrinogen was separated from unfractioned fibrinogen by ammonium sulphate precipitation. Cells were incubated with unfractioned fibrinogen, HMW-fibrinogen or fibrin. Cytokine levels in cell lysates were determined using ELISA assays. Real-time PCR was used for mRNA quantification. We found that fibrinogen significantly increased mRNA levels, and induced synthesis of the proinflammatory cytokines IL-6 and TNF- α in PBMC in a dose dependent manner. Median (25, 75 percentile) IL-6 and TNF- α concentrations were 12 (5, 40) pg/ ml and 16 (0,61) pg/ml,respectively.Median mRNA quantity was increased 12.3– (6.6, 48.6) and 1.7– (1.5, 6.5) fold for IL-6 and TNF- α compared to controls.The stimulatory effect of unfractioned fibrinogen was not significantly different from HMW-fibrinogen. Fibrinogen and fibrin were equally effective in promoting cytokine synthesis from PBMC.The results support that fibrin and fibrinogen may actively modulate the inflammatory process by inducing synthesis of proinflammatory cytokines from PBMC.

scholarly journals Neutralization of Interleukin-10 Significantly Enhances Gamma Interferon Expression in Peripheral Blood by Stimulation with Johnin Purified Protein Derivative and by Infection with Mycobacterium avium subsp. paratuberculosis in Experimentally Infected Cattle with Paratuberculosis

2004 ◽  
Vol 72 (4) ◽  
pp. 2425-2428 ◽  
Author(s):  
Joram J. Buza ◽  
Hirokazu Hikono ◽  
Yasuyuki Mori ◽  
Reiko Nagata ◽  
Sachiyo Hirayama ◽  
...  
Keyword(s):  
Mycobacterium Avium ◽  
Peripheral Blood ◽  
Mononuclear Cells ◽  
Interleukin 10 ◽  
Suppressive Effect ◽  
Gamma Interferon ◽  
Peripheral Blood Mononuclear ◽  
Purified Protein Derivative ◽  
Ifn Γ

ABSTRACT Monoclonal antibody neutralization of interleukin-10 (IL-10) increased Johnin purified protein derivative-induced whole-blood gamma interferon (IFN-γ) secretion 23-fold and also increased IFN-γ secretion ninefold following in vitro Mycobacterium avium subsp. paratuberculosis infection of peripheral blood mononuclear cells. These results demonstrate the suppressive effect of IL-10 on immune responses to M. avium subsp. paratuberculosis infection in cattle.

scholarly journals Naturally Induced Secretions of the Potato Cyst Nematode Co-stimulate the Proliferation of Both Tobacco Leaf Protoplasts and Human Peripheral Blood Mononuclear Cells

1999 ◽  
Vol 12 (10) ◽  
pp. 872-881 ◽  
Author(s):  
Aska Goverse ◽  
Jeroen Rouppe van der Voort ◽  
Charlotte Rouppe van der Voort ◽  
Annemieke Kavelaars ◽  
Geert Smant ◽  
...  
Keyword(s):  
Molecular Weight ◽  
Mononuclear Cells ◽  
Human Peripheral Blood ◽  
Plant Cells ◽  
Cyst Nematode ◽  
Potato Cyst Nematode ◽  
Low Molecular Weight ◽  
Peripheral Blood Mononuclear ◽  
Blood Mononuclear Cells ◽  
Leaf Protoplasts

Naturally induced secretions from infective juveniles of the potato cyst nematode Globodera rostochiensis co-stimulate the proliferation of tobacco leaf protoplasts in the presence of the synthetic phytohormones α-naphtha-leneacetic acid (NAA) and 6-benzylaminopurine (BAP). With the use of a protoplast-based bioassay, a low-molecular-weight peptide(s) (<3 kDa) was shown to be responsible for the observed effect. This mitogenic oligopeptide(s) is functionally dissimilar to auxin and cytokinin and, in addition, it does not change the sensitivity of the protoplasts toward these phytohormones. In combination with the mitogen phytohemagglutinin (PHA), cyst nematode secretions also co-stimulated mitogenesis in human peripheral blood mononuclear cells (PBMC). The stimulation of plant cells isolated from nontarget tissue—these nematodes normally invade the roots of potato plants—suggests the activation of a general signal transduction mechanism(s) by an oligopeptide(s) secreted by the nematode. Whether a similar oligopeptide-induced mechanism underlies human PBMC activation remains to be investigated. Reactivation of the cell cycle is a crucial event in feeding cell formation by cyst nematodes. The secretion of a mitogenic low-molecular-weight peptide(s) by infective juveniles of the potato cyst nematode could contribute to the redifferentiation of plant cells into such a feeding cell.

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