Effect of Zinc Oxide Nanoparticles on The Spleen of Adult Male Albino Rat and The Possible Protective Role of Selenium: Microanatomical and Biochemical Study.

: Thirty five adult male albino rats were used in this study. They were divided into 7 groups: Group I (Control), Group II (low dose Zno NPs group) , Group III (medium dose Zno NPs group) , Group IV (high dose Zno NPs group) , Group V (low dose Zno NPs-selenite group), Group VI (medium dose Zno NPs-selenite group), Group VII (high dose Zno NPs-selenite group): tissue of the spleen was examined histopathologically and immunohistochemical , Determination of blood cells count (number of white blood cells (WBCs), red blood corpuscles (RBCs) and platelets. Also tissue level of Hydrogen peroxide (H2O2) and paraoxonase-1(pon-1): for detection of oxidative stress estimated for all groups, PCR detection of apoptosis inducing factor (AIF), apoptotic protease activating factor-1 and caspases 3 and Zinc level in the spleen. Results of the present study showed that, compared to controls: orally intake of Zinc oxide nanoparticles (Zno NPs) cause marked changes in spleen tissues as thickening of capsule, thickening of trabeculae , destruction of white pulp and red pulp expansion especially in groups receiving zinc oxide nanoparticles only and these changes markedly decreased on groups receiving selenium as protection.


Group V (low dose Zno NPs-selenite group):
consists of 5 rats that received 100 mg/kg/day zinc oxide nanoparticles by oral gavage and 1.25 mg Se/kg/day by intraperitoneal injection (11).
Group VI (medium dose Zno NPs-selenite group): consists of 5 rats that received 200 mg/kg/day zinc oxide nanoparticles by oral gavage and 1.25 mg Se/kg/day by intraperitoneal injection (11) .
Group VII (high dose Zno NPs-selenite group): consists of 5 rats that received 300 mg/kg/day zinc oxide nanoparticles by oral gavage and 1.25 mg Se/kg/day by intraperitoneal injection (11).

1-Zinc oxide nanoparticles (Zno NPs):
Zinc oxide nanoparticles (Zno NPs) were purchased from Sigma-Aldrich Chemical Co., Egypt in the form of a bottle of 100 g that were dissolved in saline.

Sodium biselenite (Se):
Sodium biselenite (Se) was purchased from Sigma Pharmaceuticals Company, Egypt in the form of vials containing 50 g that was dissolved in saline solution.
The drugs were administered once daily for 14 days.

Scarification:
The animals were sacrificed at the end of the 1-H and E to demonstrate the histological changes (12).

3-Immunohistochemical study:
a. The anti-proliferating cell nuclear antigen (anti-PCNA) for detection of the proliferative capacity.
b. The anti-p53 factor was performed for detection of apoptosis.
After addition of ZnO NPs, the normal architecture of the spleen is distorted, group II, III, IV showed (deformed white pulp

C
The stained section of control group was characterized by strong positive brown immunoexpression of (PCNA) in most splenocytes of of the white pulp as well as in the red pulp (Fig.A), Also a weak positive brown immunoexpression of (p53) in a few numbers of cells in the RP while in the WP (Fig.D).
Results of groups II, III, IV after addition of ZnO NPs showed strong P53 immunoreaction (Fig.E) and weak PCNA (Fig.B) immunoreaction. But in groups V, VI, VII, concomitant treatment with selenium showed weak P53 immunoreaction

II) Biochemical results:
There was significant decrease in red blood cells, platelets counts, paraoxonase-1 (PON-1) and mean area percentage of PCNA in ZnO-NPs groups (group II, III, IV) compared to the control group. There was significant increase in red blood cells, platelets counts, paraoxonase-1 (PON-1) and mean area percentage of PCNA in ZnO-NPs &se groups (groups V, VI, VII).
There was significant increase in white blood cells count, Hydrogen peroxide (H2O2), %Masson's trichome Nano-ZnO, makes zinc more easily to be absorbed by the body due to small particle size, thus nano-ZnO is commonly used as a food additive .In the present work, the oral route administration was chosen as it is the main mode of exposure to ZnO NPs in humans and animals (16) The spleen is not only an organ that serves as the largest blood filter in the body, it also  Not only spleen but most of organ showed accumulation of zinc after of orally exposed ZnO-NPs in agreement with (51) demonstrate that most organs were the target of orally exposed ZnO-NPs and ZnO submicroparticles.