الفهرس 
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Abstract
Summary
Acute pancreatitis (AP) is one of the most frequent emergencies requiring hospitalization. Induction of acute pancreatitis using L-arginine is considered one of the most accepted models of AP as it produces biochemical and histological changes in the pancreas resembling those in humans.Nifedipine is used in treatment of hypertension and other cardiovascular disorders. It exerts beneficial effects in attenuating AP through its potentl anti-inflammatory and anti-oxidant properties.
The aim of this study was to investigate the possible protective effect of Nifedipine against L-arginine induced acute pancreatitis in rats.
This evaluation included the biochemical, histological, immunohistochemical and morphometric assessments.
This study was carried out on forty adult male albino rats. They were divided randomly into five groups as follows:
1. group I (Control group): Rats received 0.9% normal saline orally for one week and received two intraperitoneal injections of 1ml saline with 1 hour interval between the two doses for one time.
2. group II (Nifedipinegroup): Rats received Nifedipine orally, at a daily dose of 5mg/Kg for one week.
3. group III (L-Arginine group): Rats received two intraperitoneal injections of L-arginine Hcl at dose of 250 mg/100 gm body weight with 1 hour interval between the two doses and rats were kept for 1 week.
4. group IV (L-Arginine + Nifedipinegroup): Rats received Nifedipine after 1h of the second dose of L-arginine at a dose of 5mg/Kg orally for one week.
5. group V (Recovery group): Rats received L- arginine only at a dose of 250 mg/100 gm body weight with 1 hour interval between the two doses and rats were kept for 2 weeks.
On the last day of the experiment, each animal’s body weight was recorded. Rats were sedated with ketamine. Before the sacrifice, blood samples were drawn from the retro-orbital venous plexus of all rats. Using cervical dislocation, the animals in the control and experimental groups were sacrificed and specimen taken from the pancreas of all experimental animals.
The plan of the present study included the following items:
I. Pancreatic weight index was calculated for each group (the mean pancreatic weight as a ratio of body weight)
II. Serological Study:
Serum lipase& Serum α amylase
III. Biochemical assay of pancreatic oxidative stress and inflammatory markers:
Measurement of pancreatic: MDA, MPO, GSH, TNFα and Nrf2
IV. Histological Study:
1. Hematoxylin and Eosin (H &E) to demonstrate the histological changes.
2. Masson’s trichrome stain to demonstrate stromal changes in collagen deposition.
3.Toluidine blue stain: to detect zymogen granules in the apical part of pancreatic acinar cells to evaluate secretory function of pancreatic acinar cells.
V. Immunohistochemical Study:
1- Caspase 3 was used as an indicator of apoptosis.
2- Nuclear factor kappa B (NF-kB) was used as an indicator of inflammation.
VI. Quantitive morphometric measurement:
1. The mean area percentage of collagen stained with Masson’s Trichrome stain.
2. The mean area percentage of Caspase 3 antibody immunopositivity.
3. The mean area percentage of NF-kB antibody immunopositivity.
VII. Statistical Analysis:
The Results of the Present Work Could be Summarized as follows:
1.Pancreatic weight index
Pancreas from groups I&II (Control & Nifedipine groups) showed normal PWI whereas L-Arginine administration caused significant increase of PWI in group III &V (L-Arginine group &Recovery group). There was a significant (p < 0.001) reduction in PWI in group IV (L-Arginine + Nifedipine group) as compared to L-Arginine group & Recovery group.
2. Biochemical results
In the present study, L-Arginine group &Recovery groups recorded a significant high levels of serum α amylase and lipase compared to Control & Nifedipine groups wherase group IV significantly decreased serum α amylase and lipase levelsas compared to L-Arginine group & Recovery group.
L-Arginine & Recovery groups revealed a significant increase in pancreatic tissue levels of MDA, MPO and TNF-α, as well as a significant decrease in GSH and Nrf2 content compared to (Control group &Nifedipine group). On the other hand, treatment with Nifedipine significantly decreased pancreatic tissue levels of MDA, MPO and TNF-α, as well as a significant increased GSH and Nrf2as compared to L-Arginine group & Recovery group, recording highest improvement in all antioxidant markers.
3. Histological results
Hematoxylin and Eosin
Histological observations of control and Nifedipine groups indicated that there was presence of normal exocrine and endocrine pancreatic tissue while the histological examination of L-Arginine group showed distortion & irregular architecture of the pancreatic acini. Dilated and congested blood capillaries were noted. Some cells expressed cytoplasmic vacuolations. Inflammatory cell infiltration was also observed. The exocrine pancreas showed widened CT stroma between the pancreatic lobules and around the acini. The islets of pancreas appeared atrophied and decreased in size. On the other hand, treatment with Nifedipine showed restoration of normal pancreatic architecture with minimal infilammatory cell infiltration.
Masson’s Trichrome stain
L-Arginine group & Recovery group demonstrated significant increase of collagen deposition displayed by increased Masson’s trichrome staining intensity compared to the rats in Control & Nifedipine groups. Treatment with Nifedipine showed decrease collagen deposition as compared as compared to L-Arginine group & Recovery group.
Toluidine blue stain
Pancreatic acini and islets of langerhans of L-Arginine group &Recovery group, showed reduction of granules compared to Control & Nifedipine groups, while treatment with Nifedipine showed restoration of pancreatic granules.
4. Immunohistochemical results
Caspase 3: Expression of Caspase 3 was used to determine the apoptosis in pancreatic tissues. There is an obvious increase in brown staining in the L-Arginine & Recovery groups, compared to the control & Nifedipine groups On the other hand, L-Arginine + Nifedipine group demonstrated minimal positivity which indicates an improvement in Caspase 3 expression. Such changes in Caspase 3 immunostaining documented in the present study were verified by histomorphometric measurements of the area % of Caspase 3 immunostaining in varios groups that revealed statistically highly significant increase in the mean area percent in L-Arginin group. However, the L-Arginine+ Nifedipine group showed highly significant decrease in the mean area percent compared to L-Arginine group.
NF-kB: The expression of NF-kB in the pancreatic cells was upregulated in the L-Arginine&Recovery groups but downregulated in L-Arginine + Nifedipine group. Both Control group & Nifedipine groups showed negative immunreaction. Such changes in NF-kB immunostaining documented in the present study were verified by histomorphometric measurements of the area % of NF-kB immunostaining in varios groups that revealed statistically highly significant increase in the mean area percent in L-Arginin group. However, the L-Arginine+ Nifedipine group showed highly significant decrease in the mean area percentagecompared to L-Arginine group.