Ultrastructural variations of rat myocardium due to Walterinnesia aegytia snake envenomation.

This study was undertaken to investigate the toxic effect of Walterinnesia aegyptia venom on the ultrastructure of rat myocardium. Male albino rats were prepared for intraperitoneal injection of saline (control group) and saline solution of W.aegyptia venom (study group) at a dose of 0.04 mg animal-1. Biopsies from the left ventricle were prepared for electron microscopy after 1 h (D1 group), 2 h (D2 group), 18 h (D3 group) and 24 h (D4 group). Myocardial cells were in a state of partial to complete contraction. The D1 group showed some mitochondrial vacuoles; D2 group demonstrated more vacuolation and alterations in the form of disorganized cristae. Similar findings were depicted in D3 group. The D4 group demonstrated, in addition, dissolution of mitochondrial cristae. Myofilaments in D3 group experienced coalescence into ill-defined amorphous masses (foci of myolysis). These masses were characterized by the presence of multiple, parallel, Z-like dark bands with disorganization of the filamentous arrangement. In the D4 group, more myolytic foci were observed. This reaction was not limited to one myocyte but extended to the neighbouring ones. Mitochondrial vacuoles were mostly associated with electron dense deposits. Glycogen particles tended to decrease as the experiment proceeded from D1 to D4. These ultrastructural changes were time dependent. They would suggest a cardiotoxic action of W.aegyptia snake venom.

Ultrastructure of resting and rh-GMCSF-treated human macrophages derived from blood monocytes.

The ultrastructure of cultured blood monocyte-derived human macrophages was investigated and correlated under the effect of different doses of rh-GMCSF (dose 1 = 25 IU/ml, dose 2 = 125 IU/ml and dose 3 = 250 IU/ml). Resting macrophages showed irregular cell borders and pseudopodia pushed out in all directions. Their cytoplasm depicted rough endoplasmic reticulum and Golgi complex in the perinuclear area. Lipid globules, primary lysosomes and mitochondria were characteristically prominent. rh-GMCSF-stimulated macrophages were more voluminous and their nuclei were irregular in outline, with predominance of euochromatin over heterochromatin. The cytoplasm was overcrowded by an increasing number of organelles including lysosomes, phagolysosomes and mitochondria. Golgi complex demonstrated a wide-spread distribution along the cells, with profound membrane expansion and cisternal dilatation; especially, in cells treated with dose 2. Electron dense osmiophilic deposits (collapsed membranes) were seen in association with lipid globules, which were commonly polarized at cell peripheries. Most of these changes were dose dependent. However, cells treated with dose 3 manifested additionally well-developed centrioles, inapparent nuclear membrane, display of microfilaments and well-established adhesions. The demonstrated ultrastructural changes in rh-GMCSF-treated human macrophages indicated pronounced activation, which supports the reported clinical effect of this cytokine.

Enhancement of leishmanicidal activity of human macrophages against Leishmania major and Leishmania donovani infection using recombinant human granulocyte macrophage colony stimulating factor.

The in vitro effect of recombinant human Granulocyte Macrophage Colony Stimulating Factor (rh-GMCSF) on the leishmanicidal activity and superoxide anion productivity of macrophages derived from human blood monocytes (MOs) were investigated. MOs treated with 25, 125, or 250 U/mL of rh-GMCSF for 72 h prior to infection with leishmania parasites, manifested significant dose-dependent increase in its leishmanicidal activities against Leishmania major and Leishmania donovani parasites. The percentage of increase in leishmanicidal activity of L. major-infected MOs were 22.71, 64.34 and 81.34, respectively while in L. donovani-infected MOs, it reached 3.01, 32.28 and 74.38, respectively. Treatment of leishmania-infected MOs with rh-GMCSF (250 U/mL) for different periods of time up to 96 hours, induced a significant time-dependent reduction in the percentage of infected cells and the parasitic load (No. of amastigotes/100 MOs). After 96 h of treatment with rh-GMCSF, the percentages of reduction in the infection rates were 82.45 in L.major-infected MOs (p < 0.001) and 39.65 in L. donovani-infected cells (p < 0.01). The percentage of reduction in the parasitic load reached 90.82 (p < 0.001) and 36.6 (p < 0.05) in MOs infected with L. major and L. donovani, respectively. The priming effect of rh-GMCSF on superoxide anion production by human MOs stimulated with phorbol myristate acetate (PMA) was both dose-dependent and time-dependent. In 72 hour-old human MOs, the maximum superoxide anion release was generated by MOs primed for 45 min with 500 U/mL of rh-GMCSF. These cells produced 8.960 +/- 2.075 nmol/5 x 10(4) MOs/ 180 min as compared to 4.563 +/- 1.773 nmol/5 x 10(4) unprimed cell control/180 min (p < 0.001).

Effect of rh-GMCSF and rh-GCSF on oxygen free radical production by human neutrophils and blood monocyte-derived human macrophages.

The in vitro effect of recombinant human granulocyte-macrophage colony stimulating factor (rh-GMCSF) and recombinant human granulocyte colony stimulating factor (rh-GCSF) on oxygen free radical (OFR) generation by human neutrophils and blood monocytes derived human macrophages stimulated with phorbol myristate acetate was investigated and compared. The production of OFR by neutrophils and macrophages was time dependent, and the maximum release of OFR by neutrophils and macrophages was measured 90 and 180 min after stimulation with phorbol myristate acetate, respectively. The priming effects or rh-GMCSF and rh-GCSF on OFR production by human neutrophils and macrophages was dose dependent. The maximum generation of OFR by neutrophils occurred when primed with 1,000 U/ml of rh-GMCSF and reached 2.383 +/- 0.191 nmol/10(5) neutrophils/90 min as compared with 1.072 +/- 0.113 nmol/10(5) neutrophils/90 min in the unprimed controls. This represents a 122.20% increase in OFR generation (p < 0.001). However, the percentage of maximum increase in OFR production was 57.84 when neutrophils were primed with a concentration of 5,000 U of rh-GCSF/ml. In 72-hour-old human macrophages, much higher levels of OFR production as compared with neutrophils were measured following stimulation with phorbol myristate acetate. The maximum generation of OFR was measured in macrophages primed for 45 min with 500 U/ml of rh-GMCSF. These cells produced 8.960 +/- 2.075 nmol/5 x 10(4) macrophage/180 min as compared with 4.563 +/- 1.773 nmol/5 x 10(4) unprimed macrophages/180 min (p < 0.001). In macrophages primed with rh-GCSF, however, the maximum OFR production was induced by a dose of 5,000 U/ml and reached 6.902 +/- 1.463 nmol/5 x 10(4) macrophages/180 min as compared with 4.563 +/- 1.773 nmol/5 x 10(4) macrophages/180 min in the unprimed controls (p < 0.05). In conclusion, the priming effect of rh-GMCSF on OFR generation by human macrophages and neutrophils was more potent than that of rh-GCSF, both in the extent of augmentation and in the dose required to produce maximum OFR generation.