M.Sc Thesis

M.Sc StudentRotenberg Aviad
SubjectThe Effect of Heparanase in Sepsis and Acute Lung Injury
DepartmentDepartment of Medicine
Supervisors PROFESSOR EMERITUS Michael Krausz
PROF. Zaid A. Abassi
Full Thesis text - in Hebrew Full thesis text - Hebrew Version


BACKGROUND: The inflammatory response to sepsis or tissue injury termed systemic inflammatory response syndrome (SIRS) frequently results in acute lung injury (ALI). A central event of septic ALI is the infiltration of recruited and activated polymorphonuclear neutrophils (PMNs) into the lung tissue, initiating tissue damage and organ dysfunction.

Heparanase, an endo-β-D-glucuronidase is capable of cleaving Heparan sulfate side chains. Heparan sulfate proteoglycans are abundant components of the extracellular matrix (ECM).

We previously demonstrated that Heparanase pretreatment attenuated acute lung injury induced by endotoxin, and significantly improved the survival of affected animals. The present research aims are to examine the effect of Heparanase treatment on neutrophil accumulation in endotoxemia, which may result in decreased permeability of pulmonary endothelium, decreased levels of adhesion molecules and reduced mortality.

METHODS: Male Sprague-Dawley rats (300-350 g) were injected with increasing doses of lipopolysaccharides (LPS). After one hour, increasing doses of Heparanase were injected via the tail vein . Twenty four hours later, the rats were sacrificed, and blood withdrawn for flow cytometry, leukocyte levels and integrin distribution. Leukocytes count and protein were also determined in BALF. MPO levels, histology and neutrophil counts were obtained from lung tissue.

RESULTS: Mortality rate following of LPS with or without Heparanase treatment was 6% (n=120). In animals treated with Heparanase (without LPS), increased levels of blood monocytes were observed, and also increased CD11b integrin expression on neutrophils and monocytes, and also increased CD18 and CD29 integrins expression on neutrophils, monocytes and lymphocytes. In the Heparanase treated groups increased numbers of lung neutrophils, increased MPO enzyme activity and increased expression of CD11b/ CD18 integrins on leukocytes were observed. No statistically significant differences were observed between Heparanase treated, and the LPS untreated groups, in neutrophils and protein concentration in BALF, and the percentage of blood neutrophils, monocytes and lymphocytes. No significant differences in pH levels, Lactate, Base Excess, partial pressure of O2, and the rat's weight were observed between LPS treated and combined LPS and Heparanase treated animals.

CONCLUSION: Heparanase administration one hour after endotoxin did not alter the outcome of rats in endotoxemia. In the present investigation the most significant finding was that the increased expression of adhesion molecules CD11b/ CD18 and CD29 on leukocytes contributed to Heparanase-mediated extravasation of blood-borne cells and invasion through the pulmonary vascular endothelium. Further studies are needed in order to evaluate the anti-inflammatory effect of Heparanase in endotoxemia .