الفهرس 
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Abstract
Glycoalkaloids (GAs) are nitrogen-containing steroidal glycosides that are toxic compounds found in several vegetables, particularly in plants of the Solanaceae family, such as tomato (Lycopersicon esculentum L.) and potato (Solanum tuberosum L.). They are considered as a natural defense compound for the plants depending on their pesticidal and fungicidal influences. α-Solanine, a trisaccharide glycoalkaloids, is the main glycoalkaloid in potatoes that is biosynthesized via cholesterol pathway. In addition to α-Solanine, a chemically comparable compound named α-chaconine, they represent 95% of total glycoalkaloids in potatoes Their levels vary according to different growing, storage environmental effects that may results in significantly high content in consumable potato tubers specially the sprouts. Total glycoalkaloids reaching 100 mg per 100 g of potato meristematic tissues have been reported, while the upper accepted limit for food safety purposes is 20 mg /100 g in potatoes Generally, glycoalkaloids are known to have some beneficial effects in human health depending on the concentration where α-Solanine and other glycoalkaloid showed anti-allergic, anti-pyretic, anti-inflammatory, and antibiotic activity and lately have showed anti-cancer effects. Several cases of poisoning in humans on the other hand, have been associated with the intake of potatoes high in GA. GAs can induce gastro-intestinal (GI) and systemic effects in vivo and in vitro mainly through disruption of cell membranes and inhibition of acetylcholinesterase activity.
Diabetes mellitus is considered the most abundant metabolic disorder in humans characterized by a chronic hyperglycemic condition resulting from defects in insulin secretion / insulin action or both. Type I diabetes is caused by a lack of insulin due to the destruction of insulin-producing beta cells in the pancreas, but diabetes mellitus type II is characterized by insulin resistance rather than lack of insulin. Obesity has always been correlated to be the primary cause of type II diabetes in people who are genetically predisposed to the disease.
The aim of the present study is to prepare α-solanine extract from potatoes and to study the effect of extracted α-solanine on type I and type II diabetic rat model induced by streptozotocin.
In order to perform this aim, potatoes were collected to perform the extraction process starting with 90 kg potato yields to collected 3.5 kg buds after 3 months storage period, after extraction & lyophilization 40 gm glycoalkaloids were collected.
The extracted α-solanine was characterized using HPLC in comparison to standard purchased α-solanine, based on HPLC result the extract contain two main peaks. The first for α-solanine (RT= 5.32) which represent 45% of the extract. The second peak represent the structurally comparable α-chaconine (RT=6.65)
In vivo studies were performed to assess the effect of α-solanine on STZ induced DI & DII rat models with an oral gavage dose of (7 mg /kg.bw) for a total of 28 days after induction of diabetes.
In vivo study started with 60 male rats divided into 6 groups C, S, DI, DIS, DII & DIIS. All groups were fed with normal diet except for DII & DIIS were fed with high fat diet prior two weeks to the beginning of the experimental study.
Treatment with α-solanine extract demonstrated a significant reduction (p≤0.001) in the total body weight in S, DIS & DIIS groups if compared to C. this reduction in BW may be explained based on the effect of α-solanine on GIT affecting absorption of nutrients. Regarding relative organ weight α-solanine induced a significant reduction in both spleen and liver relative weight in DIIS compared to DII (p≤0.001) & (p≤0.05) respectively for spleen & liver, as well as a significant reduction DIS compared to DI in both spleen and liver (p≤0.05) & (p≤0.001). S itself reduced spleen & liver relative weight but non-significantly.
The FBG level was determined weekly using Accucheck® by puncturing tail tip of the animals. The FBG level showed no significant effect on S group compared to C group throughout the 4 weeks. DIS on the other hand represented aggravated hyperglycemia as compared to DI (p≤0.001) or any other group for the whole four weeks (ranging from 448.63 to 352.21 mg/dl). The result of DIIS on the other hand showed some fluctuation in hyperglycemia throughout the 4 weeks (ranging from 441.6 to 191.6). The FBG of DIIS was all the time higher but aligned with DII rats throughout the study, such elevation was statistically significant (p≤0.001). DIIS & DII showed significant elevation in FBG if compared to both S treated non-diabetic control & control group (p≤0.001). A similar fluctuation in DII STZ induced models was reported earlier and was explained based on the ability of the limited surviving β-cells in the pancreas to regenerate with time, such ability was reported to be dependent on both the dose and the age of the tested animals. Previous report of the effect of S on FBG in non-diabetic animals showed a variation between obtaining a decline in glucose level vs. elevation followed by gradual restore of blood glucose level after 24 hrs. Hypoglycemic effect was reported in both mice and rat diabetic models using extracts containing solanine but at a significantly higher administered concentration. One of the studies compared the effect of dose reporting that the anti-diabetic effect was only observed at a high dose of the extract 500 mg/kg.bw.
The resulted serum insulin level showed concordance between the hormone levels and the corresponding FBG level in most of the groups. The least detected level was in DIS group that was significantly lower than DI (p≤0.001) or any other group. The same concordance in results was observed in the DIIS compared to DII (p≤0.05). α-solanine nondiabetic control group on the other hand revealed a decline in the insulin level that was not observed to have the complementary expected elevated effect on the FBG level if compared to the control group. There’re no previous studies highlighting the relation between the level of insulin hormone in blood and α-solanine treatment.
Kidney function tests results didn’t highlight any significant changes in the Creatinine, BUN or blood serum urea in any of the included groups. Uric acid on the other hand reported a significant variation in DIS group compared to DI, where a significant increase in uric acid was reported returning the diminished values of DI to normal control level. DIIS as well, showed the same pattern with significant (p≤0.05) increase in uric acid concentration in blood compared to DII. Some studies highlighted effect of solanine on kidney parameters urea & creatinine, but none of the previous studies highlighted similar effect of uric acid in treatment with solanine.
Liver function tests results didn’t highlight any significant changes in the total protein content in any of the included groups. All groups were within the normal range in AST (50-150 IU/L) except DI & DII groups that were above the normal range. Its level was significantly higher in DI compared to the C (p≤0.05). DIS group showed a significant decline in AST activity when compared to DI (p≤0.05). A similar behavior was observed for DIIS decreasing AST in blood to the normal, but this change was not statistically significant. DI & DII groups on the other hand, reported above normal level (10-40 IU/L) in ALT, but the rest of groups were within the normal range. S administration reported the same ameliorating effect in diabetic groups, it significantly decreased ALT level in S compared to the C group. It also decreased the ALT activity in related groups DIS vs DI & DIIS vs DII (p≤0.05). Previous studies reported hepatotoxic effect of solanine demonstrating by elevation liver transaminases when a higher dose of solanine was used (50 mg/kg.bw), another study reported no effect on poultry when potato containing 4-8 g/kg solanine was used as a feeding source. Our explanation to such variation in results goes to the administration of limited solanine concentration in our study and the observed decrease in the liver organ relative weight in the solanine treated groups.
Lipid profile tests results didn’t observe any significant changes in the cholesterol, HDL, non-HDL nor LDL in any of the included groups. Despite that TG level was within the normal range (<150 mg/dl) for all groups, DI, DII & S reported higher level compared to the C (p≤0.05). DIS group showed a decline in TG activity when compared to DI. DIIS as well decreased TG when comparing to DII. Both DIS & DIIS decline was not statistically significant. Similar normalizing effect of extract containing solanine was reported in previous studies.
For the oxidative stress MDA was elevated in all groups compared to C. The highest level was detected in DIS significantly higher than DI (p≤0.05) followed by DIIS which is significantly higher than DII (p≤0.05). Finally, non-diabetic S group was higher in the oxidative stress than the C, but with non-statistical significance difference (p≤0.1331).
Regarding gene expression, both relative expression level of AKT1, TNF-α were assessed in the spleen, while CDC42 expression level was determined in pancreas. AKT downregulation was the main observation in all treated groups except for DII. Several recent research in interested the role of solanine as anti-cancer demonstrating its involving in autophagy through manipulating of AKT downstream pathways as AKT/mTor & AKT/GSK-3α. For autophagy induction AKT downregulation or deactivation-dephosphorylation may be a participating role.
TNF-α on the other hand, represents a debate regarding its participation in DM. Some studies reported an elevating TNF-α level or expression in correlation with DM, but other studies reported non-significant difference between diabetic & control groups contradicting to a third category of research articles documented a downregulation or decline level of TNF-α in diabetic models.
In pancreas CDC42 expression level was determined showing a significant downregulation in all groups compared to control. CDC42 is a recently identified regulator for pancreatic release mechanism of insulin response, as well as cellular response.
Histopathological studies were performed on sections in pancreas, spleen & liver. Regarding the pancreas, major architectural changes were observed in diabetic groups especially DI & DIS demonstrating markedly apoptotic beta cells. Apoptotic cells were less common on the other hand, in both DII & DIIS. The pancreas of S group showed no difference in comparison to C.
Liver histopathology represented mildly edematous portal tract, mildly congested portal veins in S group if compared to C. DIS showed average portal veins & portal tracts in comparison to DI which showed expanded portal tracts moderate portal inflammatory infiltrate & mildly congested portal veins. DIIS showed average hepatocytes in peri-venular areas & mild apoptosis of hepatocytes in peri-portal area in comparison to DII scattered apoptotic hepatocytes in peri-venular area & few scattered hepatocytes in peri-portal area.
Spleen histopathology represent showed no difference S if compared to C. DIS showed thick edematous capsule, atrophied lymphoid follicles & scattered apoptotic cells in peri-arteriolar area in comparison to DI showed thick capsule, peri-splenic inflammatory infiltrate, lymphoid follicles with central arterioles & average lymphocytes in peri-arteriolar area and excess ciderophages. DIIS showed average capsule, lymphoid follicles & blood sinusoids in comparison to DII thick capsule, small sized & atrophied lymphoid follicles & mildly dilated blood sinusoids.