RESISTANCE OF TRANSGENIC MICE EXPRESSING HUMAN GROUP II PHOSPHOLIPASE A2 TO E. COLI INFECTION

ABSTRACT Group II phospholipase A2 (PLA2) is a newly recognized antibacterial acute-phase protein. Recently we observed that transgenic mice expressing group II PLA2 (PLA2+ mice) were able to resist experimental Staphylococcus aureusinfection by killing the bacteria, as indicated by improved survival and by the small numbers of live bacteria in their tissues (V. J. O. Laine, D. S. Grass, and T. J. Nevalainen, J. Immunol. 162:7402–7408, 1999). To establish the role of group II PLA2 in Escherichia coli infection, the host responses of PLA2+ mice and their PLA2-deficient C57BL/6J littermates (PLA2− mice) were studied after intraperitoneal administration of E. coli. The levels of group II PLA2 in sera of PLA2+ mice increased after the administration ofE. coli, and the concentration of group II PLA2 correlated significantly with the catalytic activity of PLA2 in serum. PLA2+ mice showed lower rates of mortality and less bacterial growth in peritoneal lavage fluid, blood, and spleen and liver tissues than PLA2− mice. Unlike the observations with staphylococcal infection, serum and peritoneal lavage fluid did not inhibit the growth of E. coli in vitro. The results indicate that expression of the group II PLA2 transgene improves the host defense of mice against E. coliinfection.

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Some properties of a human group II phospholipase A2 expressed from a synthetic gene In E. coli

Biochemical Society Transactions ◽

10.1042/bst022317s ◽

1994 ◽

Vol 22 (3) ◽

pp. 317S-317S ◽

Cited By ~ 1

Author(s):

ROOHAIDA OTHMAN ◽

ANDREW WORRALL ◽

DAVID C. WILTON

Keyword(s):

Phospholipase A2 ◽

Synthetic Gene ◽

Human Group ◽

E Coli ◽

Group Ii

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The Role of Propeptide in the Refolding of Human Group IB Phospholipase A2

Acta Biochimica et Biophysica Sinica ◽

10.1093/abbs/36.9.583 ◽

2004 ◽

Vol 36 (9) ◽

pp. 583-588 ◽

Cited By ~ 1

Author(s):

Hong-Qiang Cheng ◽

Gen-Jun Xu

Keyword(s):

Light Scattering ◽

Phospholipase A2 ◽

Disulfide Bonds ◽

Human Group ◽

Terminal Sequence ◽

E Coli ◽

Mature Enzyme

Abstract Human group IB phospholipase A2 (IB-PLA2) and its zymogen (proIB-PLA2) were purified from E. coli. Refolding was carried out by diluting the denatured forms of both IB-PLA2 and proIB-PLA2 with renaturation buffer in which the disulfide bonds were completely reduced. The refolding yield of proIB-PLA2 was increased by about 50% over that of the mature enzyme. The refolding of IB-PLA2 usually produced aggregates under normal conditions, as determined by light scattering. In addition, the unfolding experiments showed that the mature enzyme was more stable than the proenzyme toward denaturants in the presence of DTT. Results suggested that the N-terminal sequence rather than its conformation of human proIB-PLA2 played an important role in the refolding process.

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Inhibition of human group II phospholipase A2 by C-reactive protein in vitro

Journal of Lipid Mediators and Cell Signalling ◽

10.1016/0929-7855(94)00037-d ◽

1995 ◽

Vol 11 (2) ◽

pp. 187-200 ◽

Cited By ~ 3

Author(s):

Peter Vadas ◽

Eva Stefanski ◽

Brigitte Grouix ◽

B.Diana Schouten ◽

Waldemar Pruzanski

Keyword(s):

Phospholipase A2 ◽

Human Group ◽

C Reactive Protein ◽

Reactive Protein ◽

Group Ii

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Human group II 14 kDa phospholipase A2 activates human platelets

Biochemical Journal ◽

10.1042/bj3270259 ◽

1997 ◽

Vol 327 (1) ◽

pp. 259-265 ◽

Cited By ~ 12

Author(s):

János POLGÁR ◽

Ruth M. KRAMER ◽

Suzane L. UM ◽

Joseph A. JAKUBOWSKI ◽

Kenneth J. CLEMETSON

Keyword(s):

Phospholipase A2 ◽

Platelet Activation ◽

Heparan Sulphate ◽

Thromboxane A2 ◽

Inhibitory Effect ◽

Human Platelets ◽

Time Dependent ◽

Human Group ◽

Group Ii ◽

Induced Aggregation

Recombinant human group II phospholipase A2 (sPLA2) added to human platelets in the low μg/ml range induced platelet activation, as demonstrated by measurement of platelet aggregation, thromboxane A2 generation and influx of intracellular free Ca2+ concentration and by detection of time-dependent tyrosine phosphorylation of platelet proteins. The presence of Ca2+ at low millimolar concentrations is a prerequisite for the activation of platelets by sPLA2. Mg2+ cannot replace Ca2+. Mg2+, given in addition to the necessary Ca2+, inhibits sPLA2-induced platelet activation. Pre-exposure to sPLA2 completely blocked the aggregating effect of a second dose of sPLA2. Albumin or indomethacin inhibited sPLA2-induced aggregation, similarly to the inhibition of arachidonic acid-induced aggregation. Platelets pre-treated with heparitinase or phosphatidylinositol-specific phospholipase C lost their ability to aggregate in response to sPLA2, although they still responded to other agonists. This suggests that a glycophosphatidylinositol-anchored platelet-membrane heparan sulphate proteoglycan is the binding site for sPLA2 on platelets. Previous reports have stated that sPLA2 is unable to activate platelets. The inhibitory effect of albumin and Mg2+, frequently used in aggregation studies, and the fact that isolated platelets lose their responsiveness to sPLA2 relatively quickly, may explain why the platelet-activating effects of sPLA2 have not been reported earlier.

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Expression of human group II PLA2 in transgenic mice results in epidermal hyperplasia in the absence of inflammatory infiltrate.

Journal of Clinical Investigation ◽

10.1172/jci118664 ◽

1996 ◽

Vol 97 (10) ◽

pp. 2233-2241 ◽

Cited By ~ 127

Author(s):

D S Grass ◽

R H Felkner ◽

M Y Chiang ◽

R E Wallace ◽

T J Nevalainen ◽

...

Keyword(s):

Transgenic Mice ◽

Inflammatory Infiltrate ◽

Human Group ◽

Epidermal Hyperplasia ◽

Group Ii

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Expression of Human Group II Phospholipase A2 in Transgenic Mice

Journal of Histochemistry & Cytochemistry ◽

10.1177/002215549704500808 ◽

1997 ◽

Vol 45 (8) ◽

pp. 1109-1119 ◽

Cited By ~ 13

Author(s):

Timo J. Nevalainen ◽

V. Jukka O. Laine ◽

David S. Grass

Keyword(s):

In Situ Hybridization ◽

Epithelial Cells ◽

Urinary Bladder ◽

Connective Tissue ◽

Transgenic Mice ◽

Phospholipase A ◽

Human Group ◽

Mouse Tissues ◽

Group Ii

Group II phospholipase A2 (PLA2) has been proposed to play an important role in inflammation and defense against bacterial infection. We investigated tissues of transgenic mice expressing the human group II PLA2 gene by immunohistochemistry using rabbit anti-human group II PLA2 antibodies, and by in situ hybridization by probing with human group II PLA2 mRNA anti-sense (test) and sense (control) riboprobes. By immunohis-tochemistry, human group II PLA2 was found in various mouse tissues and cell types including hepatocytes, proximal tubule cells of the kidney, epithelial cells of the renal pelvis, urinary bladder and ureter, granulosa cells of Graafian follicles, aortic intima and media, cartilage, epiphyseal bone, bronchial epithelial cells, and connective tissue cells in the dermis. By in situ hybridization, group II PLA2 mRNA was localized in hepatocytes, epidermal cells, dermal cells, connective tissue fibroblasts, epithelial and smooth muscle cells of the urinary bladder, and cells of Bowman's capsule. These results show that human group II PLA2 is expressed in large amounts in hepatocytes and many extrahepatic tissues of the transgenic mice. These animals provide a useful new tool for studies on the metabolism, in vivo effects, and physiological and pathological roles of phospholipase A2. (J Histochem Cytochem 45:1109–1119, 1997)

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