Received: 6 April 2021 | Revised: 26 May 2021 | Accepted: 29 May 2021 DOI: 10.1002/fsn3.2405 ORIGINAL RESEARCH Antidiabetic effects of the ethanolic extract of Allium saralicum R.M. Fritsch on streptozotocin-induced diabetes in a mice model Simin Fazelipour1 | Mahsa Hadipour Jahromy2 | Zahra Tootian3 | Nader Goodarzi4 1 Department of Anatomy, Faculty of Medicine, Tehran Medical Sciences, Islamic Azad University, Tehran, Iran 2 Herbal Pharmacology Research Center, Faculty of Medicine, Tehran Medical Sciences, Islamic Azad University, Tehran, Iran 3 Department of Basic Sciences, Faculty of Veterinary Medicine, University of Tehran, Tehran, Iran 4 Department of Basic Sciences and Pathobiology, Faculty of Veterinary Medicine, Razi University, Kermanshah, Iran Correspondence Simin Fazelipour, Department of Anatomy, Faculty of Medicine, Tehran Medical Sciences, Islamic Azad University, Tehran, Iran. Email: simin_fazelipour@yahoo.com Nader Goodarzi, Department of Basic Sciences and Pathobiology, Faculty of Veterinary Medicine, Razi University, Kermanshah, Iran. Email: n.goodarzi@razi.ac.ir Abstract Medicinal plants can protect different organs against diabetes-induced oxidative stress due to their antioxidant compounds. The present study was designed to investigate the potential of Allium saralicum R.M. Fritsch (A. saralicum) ethanolic extract to alleviate the adverse effects of streptozotocin (STZ)-induced diabetes in male BALB/c mice. Seventy male mice were randomly divided into seven groups (n = 10). Diabetes was experimentally induced by STZ (60 mg/kg bw). A. saralicum ethanolic extract with doses 5, 20, 80, and 320 mg/kg was administrated for 20 consecutive days in diabetic animals. Based on the obtained results, the untreated diabetic mice showed high blood glucose level, cholesterol, low-density lipoprotein (LDL), white blood cells count (WBC), and platelets, as well as liver enzymes, urea, and creatinine. Administration of different doses of A. saralicum extract significantly reduced blood glucose level similar to glibenclamide. Also, the levels of catalase and superoxide dismutase enzymes restored toward normal level. All hepatic and renal function parameters as well as hematological parameters were improved following treatment with A. saralicum extract particularly at high doses. Histopathological studies showed a decrease in hepatic, renal, and pancreatic damage after treatment with A. saralicum extract. The results of the present work indicate that A. saralicum ethanolic extract can attenuate diabetic hepato-renal, pancreatic, and hematological damages. KEYWORDS Allium saralicum, anemia, diabetes, liver, mice, streptozotocin 1 | I NTRO D U C TI O N resistance occurs in the body (Kayar & Agin, 2019). The main clinical complications of the diabetes mellitus are weak cutaneous wound Diabetes mellitus is a common metabolite disorder in both devel- healing, reduced fibrinolytic activity, severe chronic atherosclero- oping and developed countries (Kaczmarczyk-Sedlak et al., 2019). sis, hypertension, dyslipidemia, disturbed hematological parame- In this regard, the body ability to yield insulin reduce or insulin ters including erythrocyte aggregation, erythrocyte deformability This is an open access article under the terms of the Creative Commons Attribution License, which permits use, distribution and reproduction in any medium, provided the original work is properly cited. © 2021 The Authors. Food Science & Nutrition published by Wiley Periodicals LLC. Food Sci Nutr. 2021;00:1–12. www.foodscience-nutrition.com | 1 2 | FAZELIPOUR Et AL. hematocrit and plasma proteins, retinal failure, and renal failure with focusing on biochemical, hematological, and histopathological (Jabłońska et al., 2019; Szymusik et al., 2019). approaches. Streptozotocin (STZ), which is used for inducing diabetes, is a very toxic agent for pancreas cells especially α and β cells (Brosius et al., 2009; Michalak et al., 2020). STZ causes DNA 2 | M ATE R I A L A N D M E TH O DS inconvenience and apoptosis in α and β cells as a nitrosourea class alkylating agent (Lenzen, 2008; Tesch & Allen, 2007). In 2.1 | Plant sample collection addition to α and β cells, the liver and kidney are also sensitive to the toxicity of STZ (Rerup, 1970; Weiss, 1982), making it arduous In this research, A. saralicum was collected from the mountains of to differentiate between diabetic hepatopathy and nephropathy Kermanshah city (34.3277° N, 47.0778° E), Iran in May 2018. AS was (Tay et al., 2005). identified by the Agriculture Faculty Research Center Herbarium, Previous studies have indicated that STZ causes diabetes by Razi University, Iran (No. 2738RUH). changing the situation of antioxidant enzymes (Weiss, 1982). The results of many reports have revealed that ethnomedicinal plants by increasing the antioxidant enzymes levels have significant poten- 2.2 | Extract preparation tials for protecting of the pancreas, kidney, and liver against several toxins such as STZ (Hagh-Nazari et al., 2017; Najafi et al., 2017). A. saralicum leaves were collected in summer, then milled after dry- Some medicinal herbs have unique content of alkaloids, naphtha- ing. 220 g of leaf powder was dissolved in 100% alcohol solution quinone, tannins, triterpenes, saponins, and flavonoids (Goodarzi for 2 days. Then, the solution was filtered through paper (Whatman et al., 2016). filter paper no.42, Millipore, USA. Cat No. 1442125) and dried in Iran, as an old civilized country, has a long history of medical room temperature. Finally, 30 g of A. saralicum ethanolic extract was sciences with celebrities appreciated and acknowledged world- stored at 4℃ (Sherkatolabbasieh et al., 2017). The obtained extract wide (Goodarzi et al., 2016; Goorani et al., 2018; Sherkatolabbasieh was used for LC-Mass analysis. et al., 2017). Iranian traditional medicine is one of the main traditional medicines in the world. Previously, Satureja khuzistanica, Opuntiastrepta cantha, Silybum marianum, Ginkgo biloba, Trigonella 2.3 | Animals foenum, Ipomoea betatas, Citrullus colocynthis, Ocimum sanctum, Vaccinumarctos taphylos, Plantago ovate, Securigera Securidaca, Allium 7-week-old male BALB/c mice (n = 70) were purchased from the sativum, Cuminum cyminum, and Panax ginseng have been examined Pastor Institute, Tehran, Iran. The mice were housed in 12 hr of light– to treat diabetes in Iranian and Asian medicines (Patti et al., 2017; dark, at 24–33℃, 45%–65% relative humidity and fed with stand- Shojaii et al., 2011). ard pellet (Crude Protein: 23% Crude Fat: 3.0% Crude Fiber: 7.0% In this regard, it is predicated that Allium saralicum (A. sarali- Acid Insoluble Ash 8% Calcium: 1%–2.5% Phosphorus: 0.9% Sodium: cum) leaves have significant antidiabetic properties. The antioxi- 0.5%–1% Moisture: 12%) and water ad libitum. The protocols were dant compounds of Allium genus are Allicin [diallyl thiosulfinate], approved by the Ethics Committee of Tehran Medical Sciences, tuberoside M, thiosulfinates, S-propargyl-L-cysteine, quercetin, Islamic Azad University (Approval no: IR.IAU.PS.REC.1398.214) S-benzyl-cysteine, diosgenin, polysulfanes, diosgenin, fisetin, di- and performed completely in line with the guidelines of the Animal allylpolysulfides, onionin A, flavonoids, furostanol saponins, allyl Ethics Committee. sulfides, saponins, glycosides, diallyl tetrasulfide, and garlic oil (Goodarzi et al., 2016; Sherkatolabbasieh et al., 2017). Previously, the gastroprotective, anticancer, hepatoprotective, nephropro- 2.4 | Diabetes induction tective, anti-inflammatory, antiobesity, antidiabetic, immunoprotective, antiparasitic, neuroprotective, antifungal, antiviral, and Diabetes was induced by intraperitoneal injection of a single dose antibacterial effects of A. saralicum have been proven (Foroughi of STZ (Sigma, St. Louis, MO, USA) dissolved in citrate buffer (0.1 M et al., 2016). Antidiabetic properties of the Allium genus can be and pH 4.5), at a dose of 60 mg/kg body weight. The animals with related to the S-benzyl-cysteine, diosgenin, polysulfanes, di- fasting blood glucose more than 350 mg/dl were considered as dia- osgenin, fisetin, allicin[diallyl thiosulfinate], tuberoside M, S- betic (Hagh-Nazari et al., 2017). allylmercaptocysteine, thiosulfinates, S-propargyl-L-cysteine, Ace-AMP1, and quercetin (Patti et al., 2017). In Iranian medicine, this plant is used to treat several types of metabolite disorders such 2.5 | Experimental design as diabetes (Foroughi et al., 2016). Therefore, in the present experiment, we aimed to survey anti- One day after induction of diabetic, the animals were classified into diabetic potentials of A. saralicum ethanolic extract in a mice model the several groups (n = 10) and treated through gavage for 20 days: | FAZELIPOUR Et AL. 1. C: Healthy group treated by 0.5 ml normal saline. 3 2.10 | Histopathological assessment 2. UTD: Diabetic group treated by 0.5 ml normal saline. 3. G20: Diabetic group treated by 0.5 ml glibenclamide (20 mg/kg). In the histopathological section of the recent study, the pancreas, liver, 4. ASEE5: Diabetic group treated by 0.5 ml A. saralicum ethanolic and kidney samples were collected and investigated after preparing tis- extract (ASEE) (5 mg/kg). sue sections. The volume density of the islets and B cells, percentage 5. ASEE20: Diabetic group treated by 0.5 ml ASEE (20 mg/kg). of B cells, number of islets, and average area of islets were measured. 6. ASEE80: Diabetic group treated by 0.5 ml ASEE (80 mg/kg). In the liver sections, the enlargement and congestion in sinusoids, cen- 7. ASEE320: Diabetic group treated by 0.5 ml ASEE (320 mg/kg). tral veins, portal veins, and hepatic arteries, sinusoids hyperemia, fibrin and mononuclear cells leakage in pericentral veins and periportal zones, The dose selection was performed according to the previous studies (Sherkatolabbasieh et al., 2017). bile ducts proliferation, hepatocytes cellular and nuclear pleomorphism, eosinophilic cytoplasmic bodies and inclusion bodies in hepatocytes, hepatocytes necrosis, and liver fibrosis and cirrhosis were evaluated. In the kidney, the enlargement and congestion in glomeruli, renal 2.6 | Blood sampling veins, and renal arteries, fibrin leakage in periglomerular zone, perirenal veins, and perirenal arteries, proximal convoluted tubules and For measuring the fasting blood glucose, the blood was taken seven distal convoluted tubules, cells necrosis, glomerular and tubular at- times (1–20 days) from the tail vein and asses by a glucometer. On rophy, and renal fibrosis were assessed. day 20 of the experiment, 6 mg/kg of xylazine and 38 mg/kg of ketamine HCl were injected into the tail vein for euthanizing the animals. Then, the bloods were extracted from the hearts for biochemical 2.11 | Statistical analysis and hematological experiments. The normality of data was determined by Kolmogorov–Smirnov test and followed by one-way ANOVA test and post hoc Duncan test. 2.7 | Determination of biochemical parameters All of the statistical analyses were conducted using SPSS 22.0 (IBM SPSS Statistics for Windows, version XX) (IBM Corp.) and a p ≤ .05 The collected samples were centrifuged for 16 min at 12,000 rpm was considered significant. The values are presented as mean ± SD. and serum separated. In serum, the levels of alkaline phosphatase (ALP), alanine aminotransferase (ALT), aspartate aminotransferase (AST), albumin, total protein, conjugated bilirubin, total bilirubin, 3 | R E S U LT S creatinine, urea, cholesterol, low-density lipoprotein (LDL), and highdensity lipoprotein (HDL) were analyzed by using diagnostic kits in 3.1 | Chemical composition of ASEE Mehr laboratory, Iran. The extract yield of A. saralicum was 6.25% (18.75g), calculated on the fresh leaves of the plant. Overall, fifteen compounds such 2.8 | Determination of hematological parameters as neophytadiene, 2-hexadecene-3,7,11,15-tetramethyl, hexadecanoic acid, phytol, linolenic acid-methyl ester, hexanedioic acid, In the hematological section, the blood samples were examined by bis (2-ethylhexyl) ester, 1,4,8,11-tetraazacyclotetradecane, hexa- a hematology analyzer. The parameters including white blood cell triacontane, nonadecene, ethanol-2-tetradecyloxy, γ-tocopherol, (WBC), red blood cell (RBC), hemoglobin (Hb), packed cell volume eicosane, vitamin E, 2-phenyl-5-methylindole, and n-ethyl-1,3- (PCV), mean corpuscular volume (MCV), mean corpuscular hemo- dithioisoindoline were identified in the ASEE using GC-MS, and globin (MCH), and mean corpuscular hemoglobin concentration linolenic acid-methyl ester (24.39%), phytol (14.19%), and neophyta- (MCHC) were assessed. diene (11.6%) were the most detected compounds (Table 1) (24). 2.9 | Evaluation of the endogenous antioxidant enzymes activities 3.2 | Effect of ASEE on fasting blood glucose concentration In this research, the levels of liver and kidney antioxidant enzymes, The effect of ASEE on fasting blood glucose level in the diabetic that is, catalase (CAT) and superoxide dismutase (SOD) were meas- mice is presented in Figure 1. There was no significant change in ured according to the Mohammadi et al., (2020) and Hemmati the blood glucose level of the control mice throughout the study. et al., (2020) methods, respectively. The blood glucose levels of the untreated diabetic mice increased | 4 FAZELIPOUR Et AL. Area (%) Retention time (min) No Compound 1 Neophytadiene 2 3 4 Phytol 5 Linolenic acid, methyl esterb 6 Hexanedioic acid, bis(2-ethylhexyl) ester 1.28 29.324 7 1,4,8,11-Tetraazacyclotetradecane 1.28 30.072 8 Hexatriacontane 3.3 30.430 9 Nonadecene 5.67 32.572 11.6 18.319 2-Hexadecene, 3,7,11,15-tetramethyl 1.4 22.133 Hexadecanoic acid 6.42 23.819 14.19 25.838 24.39 26.253 b b 10 Ethanol, 2-tetradecyloxy 6 34.705 11 γ-Tocopherol 3.03 36.926 12 Eicosane 2.89 37.502 13 Vitamin E 6.14 38.295 14 2-Phenyl−5-methylindole 6.82 42.228 15 N-ethyl−1,3-dithioisoindoline 2.16 44.283 Total a a TA B L E 1 The components of ASEE analyzed by GC/MS 96.57% Expressed as percentage of the total peak area. b The dominant compounds are indicated in bold. F I G U R E 1 Fasting blood glucose levels on different days in the controls and ASEE-treated groups to approximately 520% (p ≤ .05) in a time-dependent manner. disorganization of the hepatic cords, hyperemia of sinusoids, enlarge- However, treatment of the diabetic mice with ASEE at all doses sig- ment and congestion of the central veins, portal veins, and hepatic nificantly (p ≤ .05) decreased the blood glucose levels similar to the arteries with mild hepatocellular necrosis, fibrin, and mononuclear glibenclamide-treated mice at day 20 of the experiment. ASEE ex- cells leakage. The hepatocytes of the untreated diabetic mice re- erted its maximum effect on day 20 of the experiment. vealed morphological changes such as pyknosis, karyorrhexis, chromatolysis, and cytoplasmic vacuolization. However, the liver of the ASEE-treated diabetic mice indicated significant improvement com- 3.3 | Histopathological findings pared to those of the untreated diabetic ones except the presence of a few mildly degenerated hepatocytes around the central veins The histological sections of the liver in the untreated diabetic mice and some cytoplasmic vacuoles. In addition, there was no evidence showed degenerative changes in the hepatocytes represented by of hemorrhages, inflammatory cells infiltration, or parenchymal cell | FAZELIPOUR Et AL. 5 necrosis in the livers of the ASEE320-treated diabetic mice. The liver decrease (p ≤ .05) in the untreated diabetic mice compared to of the control group had normal structure (Table 2, Figure 2). the normal control group. The volume density of pancreatic islets The kidneys of the control and ASEE-treated mice had nor- also showed a significant decline (p ≤ .05) following induction of mal structure. The proximal and distal convoluted tubules, renal diabetes. The pancreas of the diabetic mice treated with ASEE corpuscles, glomerulus, and glomerular capsule had normal archi- showed a slight increase in the size of pancreatic islets, having tecture, and in the untreated diabetic group, structural defects a few cells with hyperchromatic nucleus and regeneration of were seen in all of the above parameters. Microscopic examina- the beta cells in the center of islets. Also a regeneration process tion of the kidneys of the diabetic mice treated with ASEE320 and of pancreatic islets was more evident in ASEE-treated groups. ASEE80 did not show tubular necrosis or necrotic changes in the Although the number per square millimeter of the pancreatic is- glomerular epithelium or glomerular and vascular hemorrhages lets, the volume density of the islets, and the volume density of (Table 3, Figure 3). the beta cells in the pancreas improved following administration The effect of ASEE on histomorphometric findings of the of ASEE320, however, the percentage of beta cells, and the vol- pancreatic tissue in the diabetic mice is presented in Figure 4. ume density of the beta cells in the pancreatic islets in the ASEE- The number of pancreatic islets, volume density of the beta cells treated mice were still significantly (p ≤ .05) lower than those of as well as the percentage of the beta cells showed a significant the control group. TA B L E 2 No 1 Histopathological analysis of the liver in controls and ASEE-treated groups Liver Changes C UTD G20 ASEE5 ASEE20 ASEE80 ASEE320 Enlargement of sinusoids − +++ + + + − − 2 Enlargement of central veins − +++ + ++ ++ + − 3 Enlargement of portal veins − +++ + ++ + − − 4 Enlargement of hepatic arteries − +++ + + + − − 5 Congestion in central veins − ++++ ++ ++ ++ + + 6 Congestion in portal veins − ++++ ++ ++ ++ + − 7 Congestion in hepatic arteries − +++ ++ ++ ++ − − 8 Fibrin leakage in pericentral veins − ++ − + + − − 9 Fibrin leakage in periportal zones − +++ + ++ + + − 10 Mononuclear cell leakage in periportal zones − ++++ ++ ++ ++ + + 11 Necrosis of hepatocytes − ++++ + + + + − F I G U R E 2 The effects of ASEE on histological structure of rat's liver. The histological appearance of liver in (a) Control, (b) non-treated diabetic, (c) glibenclamide-treated, (d) diabetic +ASEE 5 mg/kg, (e) diabetic +ASEE 20 mg/kg, (f) diabetic +ASEE 80, and (g) diabetic +ASEE 320 mg/kg treated groups. The sections show normal structure in control, glibenclamid-treated and ASEE320 mg/ kg treated rats and necrotic and fibrotic changes in non-treated diabetic rats. Yellow arrows: necrotic zone, White arrows: vascular congestion (H&E, 100×) (a) (b) (c) (d) (e) (f) (g) | 6 TA B L E 3 FAZELIPOUR Et AL. Histopathological analysis of kidney in controls and ASEE-treated groups No Kidney Changes C UTD G20 ASEE5 ASEE20 ASEE80 ASEE320 1 Enlargement of glomeruli − ++ − − − − − 2 Enlargement of renal vein − ++ − + − − − 3 Enlargement of renal artery − ++ − − − − − 4 Congestion in renal vein − +++ + + + − − 5 Congestion in renal artery − ++ − − − − − 8 Fibrin leakage in perirenal vein − ++ − + − − − 6 Fibrin leakage in perirenal artery − + − − − − − 7 Necrosis of proximal convoluted tubule cells − ++ − + + − − 8 Necrosis of distal convoluted tubule cells − ++ − − − − − 9 Glomerular and tubular atrophy − + − − − − − (a) (b) (c) (d) (e) (f) F I G U R E 3 The effects of ASEE on histological structure of rat's kidney. The histological appearance of liber in (a) Control, (b) non-treated diabetic, (c) glibenclamide-treated, (d) diabetic +ASEE 5 mg/kg, (e) diabetic +ASEE 20 mg/kg, (f) diabetic +ASEE 80 and (g) diabetic +ASEE 320 mg/kg treated groups. The sections show normal structure in control, glibenclamid-treated and ASEE320 mg/kg treated rats. Yellow circle: inflammatory cells infiltration, Blue circles: necrotic glomeruli, Black stars: necrotic tubules (H&E, 100×) (H&E, 100×) (g) 3.4 | Effect of ASEE on liver biochemical parameters creatinine levels and decreased CAT and SOD levels significantly The estimated values of the liver enzymes are presented in Figures 5–7. could significantly (p ≤ .05) ameliorate the above parameters. There STZ-induced hepatotoxicity increased ALP, AST, ALT, cholesterol, LDL, was no significant difference in the above-mentioned parameters total, and conjugated bilirubin and decreased HDL, SOD, CAT, total pro- (p ≤ .05) between ASEE320 and control groups. (p ≤ .05) compared to the control group. Different doses of ASEE tein, and albumin significantly (p ≤ .05) as compared to the control group. Several doses of ASEE and glibenclamide could significantly (p ≤ .05) decrease the raised levels of ALP, AST, ALT, cholesterol, LDL, total and 3.6 | Effect of ASEE on hematological parameters conjugated bilirubin and increased HDL, SOD, CAT, total protein, and albumin significantly (p ≤ .05) as compared to the untreated group. The number of WBC and platelet and percentage of eosinophils and basophils significantly (p ≤ .05) increased in the untreated diabetic 3.5 | Effect of ASEE on kidney biochemical parameters mice. Also, the levels of lymphocytes, monocytes, RBC, MCV, Hb, MCH, MCHC, and PCV significantly (p ≤ .05) reduced in the untreated diabetic group. Treatment with ASEE significantly (p ≤ .05) ameliorated the above parameters. There were no significant dif- The estimated values of the kidney biochemical parameters are ferences in hematological parameters (p ≤ .05) between ASEE5, depicted in Figure 8. STZ-induced diabetes increased urea and ASEE20, and glibenclamide groups. Also there were no significant | FAZELIPOUR Et AL. 7 F I G U R E 4 (a) VDI, and VDBCP and (b) NI, VDCI, and PBC values in the controls and ASEE-treated groups. NI (Number of islets (per mm2)), VDCI (B cells volume density in islets), PBC (B cells percent), VDI (Islets volume density), and VDBCP (B cells volume density in pancreas) FIGURE 5 groups (a) The serum levels of ALP, AST, ALT (IU/L), and (b) cholesterol, LDL, and HDL (mmol/L) in the controls and ASEE-treated differences (p ≤ .05) in the above factors (except for platelet and Hb or increase in glucose uptake by peripheral tissues (Gupta et al., 2012; levels) between ASEE320 and control groups (Figures 9–11). Hamden et al., 2001; Porchezhian et al., 2000). In addition, herbal extracts may have stimulatory effects on the remaining beta cells and more insulin production. Various studies have shown that the 4 | D I S CU S S I O N administration of plant extracts in laboratory diabetic animals can be effective in the reconstruction and repairment of the beta cells and The present study investigates the efficacy of ethanolic extract of Langerhans islands. Beta cells also have a remarkable potential for Allium saralicum (A. saralicum) on streptozotocin-induced diabetes in self-renewing in the early stages of diabetes (Cumaoğlu et al., 2011; male mice from various histopathological, hematological, and bio- Pepato et al., 2004). Therefore, by default, it can be assumed that the chemical aspects. ethanolic extract of A. saralicum is responsible for the production and The obtained results showed that the ethanolic extract of A. sarali- secretion of insulin from recombinant beta cells in the pancreas. Based cum could significantly reduce the blood glucose level in STZ-induced on the histopathological findings in this study, the volume density of diabetes. Such hypoglycemic effects of medicinal plants can be at- the beta cells and also the number and size of the Langerhans islands tributed to decrease in the rate and amount of intestinal absorption showed a significant improvement in the diabetic mice treated with 8 | FAZELIPOUR Et AL. FIGURE 6 (a) Total protein and albumin levels and (b) Total bilirubin and conjugated bilirubin in the controls and ASEE-treated groups F I G U R E 7 The serum levels of liver SOD and CAT in the controls and ASEEtreated groups ethanolic extract of A. saralicum. These changes can explain declined ethanolic extract of A. saralicum has been able to improve the defect blood glucose levels in these groups. In addition, many researchers metabolism of the fatty acids in the streptozotocin-induced diabetic have suggested that the antidiabetic effects of some of the natural mice. Increased lipid decomposition and the release of free fatty acids extracts can be attributed to their insulin-like effects, which enable from peripheral tissues are other mechanisms for increasing the lip- them to decrease the blood glucose levels and serum lipids by con- ids profile in diabetes (Chaiyasut et al., 2011). Previous studies have trolling insulin (Shen et al., 2000; Zangeneh et al., 2018). shown that some compounds, especially saponins and steroids, exert Dyslipidemia is one of the complications of hyperglycemia antihyperlipidemic effects by preventing intestinal absorption of lip- (Adeneye et al., 2010). This work showed that cholesterol and LDL ids and also by preventing the activity of lipase enzymes (Hamden levels decreased and HDL levels enhanced in the diabetic mice treated et al., 2001). The increase in the liver enzymes in diabetic mice may with ethanolic extract of A. saralicum. These results suggest that the be due to diabetes-induced hepatic injuries (Rodrigues et al., 2010). | FAZELIPOUR Et AL. FIGURE 8 (a) The serum levels of urea, creatinine, and (b) kidney SOD and CAT levels in the controls and ASEE-treated groups FIGURE 9 The number of (a) WBC, (b) platelet, and (c) RBC in the controls and ASEE-treated groups 9 The findings of the current work confirmed the signs of liver damage, chromatography/spectrometry, linolenic acid is the most effective such as dilation and congestion of the sinusoids, hepatic arteries, and ingredient found in A. saralicum extract. According to the previous veins. Treatment with different doses of A. saralicum ethanolic extract studies, this fatty acid known as herbal omega-3 has antioxidant and reduced the serum levels of transaminases and also improved the anti-inflammatory potentials and was used for treating various dis- histopathologic alterations in the liver of the streptozotocin-induced eases such as experimental colitis, vascular thrombosis, osteoporosis, diabetic animals. Also, diabetes can increase the bilirubin levels di- and myocardial infarction. This fatty acid has an excellent inhibitory rectly by damaging the bile ducts or by releasing from the muscles effect on NO and iNOS production. Also, the antioxidant effects of (Gaamoussi et al., 2010). The obtained results showed that the conju- linolenic acid can be attributed to its ability to regulate the expres- gated and total bilirubin levels restored toward the normal levels after sion of TNF-α as well as inflammatory interleukins (Sherkatolabbasieh treating with high doses of ethanolic extract of A. saralicum. et al., 2017). Moreover, other compounds in A. saralicum extract, such It has been well established that stress oxidative plays a pivotal as phytol, neofitadine, and vitamin E, are potent antioxidant and anti- role in the pathogenesis of diabetes and vascular complications. inflammatory agents (Ren & Chung, 2007). Hematological indices Streptozotocin can increases reactive oxygen species (ROS) produc- were another parameter studied in this study. In general, the rela- tion and damages to the pancreas, leading to increased blood glucose tionship between diabetes and anemia has been fully documented in level. These molecules are exacerbating factors in cellular injury, in- previous studies (Mehdi & Toto, 2009; Weiss & Goodnough, 2005). flammation, cardiovascular diseases, and aging process. Therefore, Several mechanisms can be considered for anemia associated with di- antioxidants play a significant role in reducing diabetes complications abetes. Ferraro et al., (2011) in a study showed that diabetes affects (Rahimi et al., 2005; Tchinda et al., 2008). Based on the results of gas the bone marrow cells and changes the microanatomy and physiology 10 | FIGURE 10 FAZELIPOUR Et AL. (a) Lymphocytes, neutrophils, monocytes, and (b) eosinophils and basophils percent in the controls and ASEE-treated groups F I G U R E 1 1 The values of (a) PCV, (b) MCV, (c) MCHC, and Hb and (d) MCH in the controls and ASEE-treated groups of the bone marrow stem cells. In addition, it seems that one of the decreased significantly. Stookey et al., (2007) have shown that strep- causes of diabetes mellitus-induced anemia is the glycosylation of the tozotocin reduces the synthesis of MCH and MCHC, which indicates a plasma membrane of the red blood cells. So that, hyperglycemia and defect in hemoglobin synthesis and a defect in osmotic pressure con- protein oxidation lead to increased lipid peroxidation and ultimately trol and osmolality of the plasma. Treatment with A. saralicum extract, diminish the fluidity and flexibility of the cell membrane and hemolysis especially at high dose (320 mg/kg), improved the above-mentioned of the red blood cells can be occurred (Kumar, 2012; Turk et al., 2002; parameters. Peelman et al., (2004) suggested that leptin and its re- Watala & Winocour, 1992). In this study, the number of white blood ceptor are responsible for hemopoiesis. Ohshita et al., (2004) showed cells and platelets increased in the untreated diabetic animals, and the that white blood cell count is associated with some diseases, including number of red blood cells, hemoglobin, MCV, MCH, MCHC, and PCV insulin resistance and diabetes (Pertynska-Marczewska et al., 2004; | FAZELIPOUR Et AL. Shurtz-Swirski et al., 2001). The findings indicated that the A. saralicum ethanolic extract, in addition to improving the reduction of red blood cell count and related parameters, also increased leukocyte indices. Since oxidative stress is the main risk factor of hemolysis of the red blood cells and anemia due to diabetes, the improvement of hematological indices can be attributed to the antioxidant properties of A. saralicum extract on controlling catalase and superoxide dismutase enzymes. 5 | CO N C LU S I O N In conclusion, according to the present results, it seems that the ethanolic extract of A. saralicum due to its antioxidant compounds such as linolenic acid, phytol, and neofitadine can improve hyperglycemia caused by diabetes, and attenuate renal, hepatic, and pancreatic complications. Also, this extract can prevent of anemia and blood disorders following diabetes by controlling the hematological parameters. These findings can be served as a light to formulate a suitable food supplement for alleviating diabetes complications in future. AC K N OW L E D G M E N T S The authors would like to thank Tehran Medical Sciences, Islamic Azad University for their financial support. C O N FL I C T O F I N T E R E S T The authors declare that they do not have any conflict of interest. AU T H O R C O N T R I B U T I O N S Simin Fazelipour: Conceptualization (equal). Mahsa Hadipour Jahromi: Investigation (equal). Zahra Tootian: Writing-original draft (equal). Nader Goodarzi: Software (equal); Visualization (equal). E T H I C A L A P P R OVA L This study was approved by the Institutional Review Board of Tehran Medical Sciences, Islamic Azad University (Approval no: IR.IAU. PS.REC.1398.214). DATA AVA I L A B I L I T Y S TAT E M E N T The data that support the findings of this study are available on request from the corresponding author. The data are not publicly available due to privacy or ethical restrictions. ORCID Nader Goodarzi https://orcid.org/0000-0003-4704-6076 REFERENCES Adeneye, A. A., Adeyemi, O. O., & Agbaje, E. O. (2010). Anti-obesity and antihyperlipidaemic effect of Hunteria umbellata seed extract in experimental hyperlipidaemia. 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