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Relationships between the structure and function of lipopolysaccharide chemotypes with regard to their effects on the
C Pugliese1, M D LaSalle, V A DeBari
1Department of Biology, Seton Hall University, South Orange, New Jersey.
Abstract:
Polymorphonuclear neutrophils (PMN), harvested from healthy volunteers, were challenged with endotoxins from Salmonella minnesota smooth-strain and rough-strain mutants (Ra, Rb2, RcP-, Rd1P- and Re) as well as with lipid A in an effort to determine the relationship between lipopolysaccharide (LPS) structure and its ability to affect PMN chemotaxis (CT), random migration (RM) and luminol-dependent chemiluminescence (CL). In the presence of untreated autologous serum (UAS), as has been previously demonstrated for CL, CT in a modified Boyden chamber is stimulated but no significant differences among the chemotypes were observed. In the presence of heat inactivated autologous serum (IAS), significant variations among the LPS chemotypes occurred. Also noteworthy was the observation that, although the total number of cells migrating across the membrane was similar for both UAS and IAS, UAS promoted PMN migration beyond the membrane into the chamber, as opposed to IAS which caused the cells to remain adherent to the distal face of the membrane. In the absence of serum, RM was inhibited in a dose-dependent fashion by LPS from lipid A and the rough-strain mutants, with the degree of inhibition being progressively greater with increasing molecular complexity of the chemotype LPS. The smooth-strain LPS exerted an intermediate effect. In the case of CL, lipid A was the most potent stimulus, with the response decreasing as molecular complexity increased up to RcP-; the remaining core mutant LPS provided slightly greater responses. The LPS from the smooth-strain promoted a response similar to the response to Ra LPS. Based on these data, it is concluded that there are qualitative as well as quantitative effects of the carbohydrate moieties of LPS. These data also suggest that while LPS may provoke active migration in the circulation during endotoxemia, infection localized in tissue might bring about an inhibition of PMN migration while allowing the PMN to mount an oxidative response.
Insights
Lipopolysaccharide (LPS) structure influences polymorphonuclear neutrophil (PMN) functions like migration and chemiluminescence, with serum presence altering these effects. Different LPS structures impact PMN activity, suggesting varied roles in endotoxemia and localized infections.
Area of Science:
- Immunology
- Microbiology
- Biochemistry
Background:
- Lipopolysaccharide (LPS) is a major component of Gram-negative bacteria.
- LPS structure varies significantly among different bacterial strains and mutants.
- Polymorphonuclear neutrophils (PMNs) are critical immune cells involved in pathogen clearance.
Purpose of the Study:
- To investigate the relationship between lipopolysaccharide (LPS) structure and its effects on PMN chemotaxis (CT), random migration (RM), and chemiluminescence (CL).
- To determine how serum presence and inactivation affect LPS-mediated PMN responses.
- To elucidate the role of LPS carbohydrate moieties in modulating PMN functional activities.
Main Methods:
- Harvesting PMNs from healthy volunteers.
- Challenging PMNs with various Salmonella minnesota LPS chemotypes (smooth and rough mutants) and lipid A.
- Assessing PMN chemotaxis using a modified Boyden chamber, random migration, and luminol-dependent chemiluminescence in the presence and absence of autologous serum (untreated and heat-inactivated).
Main Results:
- In untreated autologous serum (UAS), LPS stimulated PMN chemotaxis, but no significant differences were observed among LPS chemotypes.
- In heat-inactivated autologous serum (IAS), significant variations in PMN responses occurred across different LPS chemotypes.
- Serum presence influenced PMN migration patterns: UAS promoted migration into the chamber, while IAS led to adherence to the membrane.
- In serum-free conditions, LPS and lipid A inhibited random migration in a dose-dependent manner, with increasing inhibition correlating with LPS molecular complexity.
- Lipid A was the most potent stimulus for chemiluminescence, with activity decreasing as LPS molecular complexity increased.
- Smooth-strain LPS showed intermediate effects on migration and chemiluminescence.
Conclusions:
- LPS structure, particularly its carbohydrate moieties, exerts both qualitative and quantitative effects on PMN functions.
- Serum plays a crucial role in modulating LPS-induced PMN responses.
- These findings suggest LPS may induce active PMN migration in circulation during endotoxemia but inhibit migration in localized infections while still eliciting an oxidative response.