THE LAST PLANNER® SYSTEM PATH CLEARING APPROACH IN ACTION: A CASE STUDY

The “Last Planner® System” (LPS) is commonly viewed as the foundation of Lean Project Delivery. It is increasingly used in certain parts of the globe. However, LPS implementation often fades off due to issues reported at organisational, project and external levels. The LPS Path Clearing Approach (PCA) offers an antidote to these issues. The goal of this paper is to outline how the LPS-PCA helped restart a stalled implementation of the LPS through a “shallow and wide” organisational approach rather than a more traditional “narrow and deep” project approach. The LPS-PCA in action is documented within an on-going UK case study organisation. Action and covert research methods were used to introduce LPS principles, thinking and language without attributing them to LPS in response to resistance to the actual LPS. The 15 step actions within the LPS-PCA are expanded from a past, current and future state perspective. The study found that the LPS-PCA’s 15 step actions were useful as a benchmark to continuously remove constraints that blocked the implementation of the LPS. In summary, the use of the LPSPCA is recommended before, during and after organisations engage with LPS Consultants if organisations are serious about sustaining the implementation of the LPS

Characterization of the Lipopolysaccharide from a \u3cem\u3eRhizobium phaseoli\u3c/em\u3e Mutant that is Defective in Infection Thread Development

The lipopolysaccharide (LPS) from a Rhizobium phaseoli mutant, CE109, was isolated and compared with that of its wild-type parent, CE3. A previous report has shown that the mutant is defective in infection thread development, and sodium dodecyl sulfate-polyacrylamide gel electrophoresis shows that it has an altered LPS (K. D. Noel, K. A. VandenBosch, and B. Kulpaca, J. Bacteriol. 168:1392-1462, 1986). Mild acid hydrolysis of the CE3 LPS released a polysaccharide and an oligosaccharide, PS1 and PS2, respectively. Mild acid hydrolysis of CE109 LPS released only an oligosaccharide. Chemical and immunochemical analyses showed that CE3-PS1 is the antigenic O chain of this strain and that CE109 LPS does not contain any of the major sugar components of CE3-PS1. CE109 oligosaccharide was identical in composition to CE3-PS2. The lipid A\u27s from both strains were very similar in composition, with only minor quantitative variations. Sodium dodecyl sulfate-polyacrylamide gel electrophoresis of CE3 and CE109 LPSs showed that CE3 LPS separated into two bands, LPS I and LPS II, while CE109 had two bands which migrated to positions similar to that of LPS II. Immunoblotting with anti-CE3 antiserum showed that LPS I contains the antigenic O chain of CE3, PS1. Anti-CE109 antiserum interacted strongly with both CE109 LPS bands and CE3 LPS II and interacted weakly with CE3 LPS I. Mild-acid hydrolysis of CE3 LPS I, extracted from the polyacrylamide gel, showed that it contained both PS1 and PS2. The results in this report showed that CE109 LPS consists of only the lipid A core and is missing the antigenic O chain

Lipopolysaccharide is a frequent contaminant of plasmid DNA preparations and can be toxic to primary cells in the presence of adenovirus

Endotoxin (lipopolysaccharide, LPS) is commonly found as a contaminant in plasmid DNA preparations. We demonstrate here that the quantities of LPS typically contaminating DNA preparations can generate a toxicity to primary cells (primary human skin fibroblasts, primary human melanoma cells) in the presence of entry-competent adenovirus particles. Toxicity can be observed with as little as 100 ng/ml free LPS or 100 pg/ml LPS when the LPS is assembled into polylysine/adenovirus complexes. Simple and effective methods of removing the contaminating LPS using either a polymyxin B resin or Triton X-114 extraction are described. Treatment of DNA samples to remove LPS eliminates the toxicity to primary cells

\u3cem\u3eRhizobium leguminosarum\u3c/em\u3e CFN42 Genetic Regions Encoding Lipopolysaccharide Structures Essential for Complete Nodule Development on Bean Plants

Eight symbiotic mutants defective in lipopolysaccharide (LPS) synthesis were isolated from Rhizobium leguminosarum biovar phaseoli CFN42. These eight strains elicited small white nodules lacking infected cells when inoculated onto bean plants. The mutants had undetectable or greatly diminished amounts of the complete LPS (LPS I), whereas amounts of an LPS lacking the O antigen (LPS II) greatly increased. Apparent LPS bands that migrated between LPS I and LPS II on sodium dodecyl sulfate-polyacrylamide gels were detected in extracts of some of the mutants. The mutant strains were complemented to wild-type LPS I content and antigenicity by DNA from a cosmid library of the wild-type genome. Most of the mutations were clustered in two genetic regions; one mutation was located in a third region. Strains complemented by DNA from two of these regions produced healthy nitrogen-fixing nodules. Strains complemented to wild-type LPS content by the other genetic region induced nodules that exhibited little or no nitrogenase activity, although nodule development was obviously enhanced by the presence of this DNA. The results support the idea that complete LPS structures, in normal amounts, are necessary for infection thread development in bean plants

Expression of \u3cem\u3eRhizobium leguminosarum\u3c/em\u3e CFN42 Genes for Lipopolysaccharide in Strains Derived from Different \u3cem\u3eR. leguminosarum\u3c/em\u3e Soil Isolates

Two mutant derivatives of Rhizobium leguminosarum ANU843 defective in lipopolysaccharide (LPS) were isolated. The LPS of both mutants lacked O antigen and some sugar residues of the LPS core oligosaccharides. Genetic regions previously cloned from another Rhizobium leguminosarum wild-type isolate, strain CFN42, were used to complement these mutants. One mutant was complemented to give LPS that was apparently identical to the LPS of strain ANU843 in antigenicity, electrophoretic mobility, and sugar composition. The other mutant was complemented by a second CFN42 lps genetic region. In this case the resulting LPS contained O-antigen sugars characteristic of donor strain CFN42 and reacted weakly with antiserum against CFN42 cells, but did not react detectably with antiserum against ANU843 cells. Therefore, one of the CFN42 lps genetic regions specifies a function that is conserved between the two R. leguminosarum wild-type isolates, whereas the other region, at least in part, specifies a strain-specific LPS structure. Transfer of these two genetic regions into wild-type strains derived from R. leguminosarum ANU843 and 128C53 gave results consistent with this conclusion. The mutants derived from strain ANU843 elicited incompletely developed clover nodules that exhibited low bacterial populations and very low nitrogenase activity. Both mutants elicited normally developed, nitrogen-fixing clover nodules when they carried CFN42 lps DNA that permitted synthesis of O-antigen-containing LPS, regardless of whether the O antigen was the one originally made by strain ANU843

Lipopolysaccharide (LPS)-binding protein is carried on lipoproteins and acts as a cofactor in the neutralization of LPS.

Lipoproteins isolated from normal human plasma can bind and neutralize bacterial lipopolysaccharide (LPS) and may represent an important mechanism in host defense against gram-negative septic shock. Recent studies have shown that experimentally elevating the levels of circulating high-density lipoproteins (HDL) provides protection against death in animal models of endotoxic shock. We sought to define the components of HDL that are required for neutralization of LPS. To accomplish this we have studied the functional neutralization of LPS by native and reconstituted HDL using a rapid assay that measures the CD14-dependent activation of leukocyte integrins on human neutrophils. We report here that reconstituted HDL particles (R-HDL), prepared from purified apolipoprotein A-I (apoA-I) combined with phospholipid and free cholesterol, are not sufficient to neutralize the biologic activity of LPS. However, addition of recombinant LPS binding protein (LBP), a protein known to transfer LPS to CD14 and enhance responses of cells to LPS, enabled prompt binding and neutralization of LPS by R-HDL. Thus, LBP appears capable of transferring LPS not only to CD14 but also to lipoprotein particles. In contrast with R-HDL, apoA-I containing lipoproteins (LpA-I) isolated from plasma by selected affinity immunosorption (SAIS) on an anti-apoA-I column, neutralized LPS without addition of exogenous LBP. Several lines of evidence demonstrated that LBP is a constituent of LpA-I in plasma. Passage of plasma over an anti-apoA-I column removed more than 99% of the LBP detectable by ELISA, whereas 31% of the LBP was recovered by elution of the column. Similarly, the ability of plasma to enable activation of neutrophils by LPS (LBP/Septin activity) was depleted and recovered by the same process. Furthermore, an immobilized anti-LBP monoclonal antibody coprecipitated apoA-I. The results described here suggest that in addition to its ability to transfer LPS to CD14, LBP may also transfer LPS to lipoproteins. Since LBP appears to be physically associated with lipoproteins in plasma, it is positioned to play an important role in the neutralization of LPS

Freedom of Information

OBJECTIVE: It has previously been shown that a combination of inhaled nitric oxide (iNO) and intravenous (IV) steroid attenuates endotoxin-induced organ damage in a 6-hour porcine endotoxemia model. We aimed to further explore these effects in a 30-hour model with attention to clinically important variables. DESIGN: Randomized controlled trial. SETTING: University animal laboratory. SUBJECTS: Domestic piglets (n = 30). INTERVENTIONS: Animals were randomized into 5 groups (n = 6 each): 1) Controls, 2) LPS-only (endotoxin/lipopolysaccharide (LPS) infusion), 3) LPS + iNO, 4) LPS + IV steroid, 5) LPS + iNO + IV steroid. MEASUREMENTS AND MAIN RESULTS: Exposure to LPS temporarily increased pulmonary artery mean pressure and impeded renal function with elevated serum creatinine and acidosis compared to a control group over the 30-hour study period. Double treatment with both iNO and IV steroid tended to blunt the deterioration in renal function, although the only significant effect was on Base Excess (p = 0.045). None of the LPS + iNO + IV steroid treated animals died during the study period, whereas one animal died in each of the other LPS-infused groups. CONCLUSIONS: This study suggests that combined early therapy with iNO and IV steroid is associated with partial protection of kidney function after 30 hours of experimental LPS infusion

Localization of lipopolysaccharide from Escherichia Coli into human atherosclerotic plaque

Experimental studies showed that gut-derived lipopolysaccharide (LPS) is pro-atherogenic, however, its relationship with human atherosclerosis is still to be defined. We investigate if gut-derived LPS from Escherichia Coli localizes in human carotid plaque and its potential role as pro-inflammatory molecule in the atherosclerotic lesion. LPS from Escherichia Coli and Toll-like receptor 4 (TLR4) were studied in specimens from carotid and thyroid arteries of 10 patients undergoing endarterectomy and 15 controls matched for demographic and clinical characteristics. Blood LPS were significantly higher in patients compared to controls. Immunochemistry analysis revealed positivity for antibodies against LPS and TLR4 coincidentally with positivity for CD68 only in the atherosclerotic plaque of carotid arteries but not in thyroid arteries; the positivity for LPS and TLR4 was greater in the area with activated macrophages. LPS concentration similar to that detected in atherosclerotic plaque resulted in a dose-dependent TLR4-mediated Nox2 up-regulation by human monocytes. These data provide the first evidence that LPS from Escherichia Coli localizes in human plaque and may contribute to atherosclerotic damage via TLR4-mediated oxidative stress