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Journal of Diabetes and Metabolic Disorders logoLink to Journal of Diabetes and Metabolic Disorders
. 2021 Sep 23;20(2):1573–1581. doi: 10.1007/s40200-021-00904-1

Investigating the effect of ethanolic extract of Commiphora myrrha (Nees) Engl. gum-resin against hepatorenal injury in diabetic rats

Mohammadmehdi Hassanzadeh-Taheri 1, Mojtaba Salimi 1, Khadijeh Vazifeshenas-Darmiyan 2, Mahtab Mohammadifard 3, Mehran Hosseini 1,✉
PMCID: PMC8630135  PMID: 34900809

Abstract

Purpose

Management of hepatorenal complications in diabetic patients is still a challenge for clinicians. The study aimed to investigate the impacts of ethanolic extract of Commiphora myrrha (Nees) Engl.oleo-gum-resin (EEM) against hepatorenal injury in diabetic rats.

Methods

Diabetes was induced by an intraperitoneal (i.p.) injection of streptozotocin (55 mg/kg) in adult male Wistar rats (n = 40); whereas, normal control rats (NC, n = 8) were treated with vehicle solution (citrate buffer, i.p.). Diabetic animals were gavaged with 500 mg/kg of metformin (MET500) and different doses of EEM (100, 300, and 500 mg/kg) once daily for 28 days. Diabetic model (DM) and NC groups were treated with normal saline. Various parameters like fasting blood glucose (FBG), plasma insulin, aspartate transaminase (AST), alanine transaminase (ALT), creatinine (Cr), urea, 24-h urine total protein (UTP), urine volume, and hepatorenal histopathology were assessed at the end of the study.

Results

Compared to the NC group, diabetic rats showed marked elevations in FBG, AST, ALT, urea, Cr, UTP, urine volume, and a significant reduction in insulin. Diabetic animals also exhibited severe histopathological alterations in liver and kidney tissues. The EEM treatment could not influence the biochemical and pathological alterations. Treatment with EEM at the dose of 300 mg/kg could slightly ameliorate some pathological alterations (fatty changes and tubular congestion) in hepatic and renal tissues.

Conclusions

These findings demonstrated that EEM treatment at doses up to 500 mg/kg could not effectively slow down the pathological process of hepatorenal damage in diabetic rats.

Keywords: Diabetes, Diabetic Nephropathies, Liver diseases, Myrrh resin

Introduction

Diabetes is among the top 10 causes of death in the world. In the last decade, its prevalence has been increased, and it is estimated that the number of people with diabetes will reach 640 million by 2050 [1]. Diabetes refers to a group of disorders that shares glucose intolerance in common. It is characterized by episodes of hyperglycemia and glucose intolerance occurring as a result of deficiencies in insulin action/secretion [2]. Elevated blood glucose levels can lead to long-term problems, including macrovascular complications associated with cardiovascular disease and microvascular complications such as retinopathy and nephropathy. In addition to these pathological conditions, diabetes can affect all body systems, including the liver [3]. Diabetic patients are at an increased risk of developing a spectrum of liver disorders [4]. Furthermore, another common complication that may occur in those with diabetes is diabetic nephropathy [5].

Over the last two decades, several hypoglycemic agents have been introduced [6]. Despite the numerous pharmacological approaches for treating diabetes that have become available, none have shown ideal results [7]. On the other hand, there has been an upsurge in the global use of herbal medicines. It has been supposed that about three-quarter of the world’s population uses complementary medicine [8]. Many diabetic patients prefer to use medicinal plants alone or in combination with conventional therapies. The main reasons for using medicinal plants are the patients’ concerns about synthetic drugs’ efficacy and possible side effects [9]. In addition to anti-diabetic effects, some herbs also have beneficial effects on diabetic complications such as liver and kidney damage [10–13].

Some medicinal plants and natural products are well-known and more accessible for a wide range of people. These compounds are used frequently for several medicinal purposes. Commiphora myrrha (Nees) Engl. oleo-gum-resin (myrrh) is one of the most famous natural products [14]. In Persian medicine, myrrh has several applications. It is used in the preparation of Aloe-based purgative medications, and also it is considered a valuable natural product for wound healing, inflammation, and management of painful swellings [15, 16]. Previous studies revealed that myrrh has anti-inflammatory, anti-tumor, antiseptic, and anti-parasitic activities [14, 17]. Moreover, there is evidence supporting the anti-diabetic potential of myrrh on type-2 diabetes [18]. Recently, we have reported that ethanolic extract of myrrh could attenuate reproductive impairments in diabetic rats [19].

There are now some remained unfolded aspects such as its effects on diabetic complications like nephropathy and liver disorders. Hence, this study was performed to evaluate the probable effects of ethanolic extract of Commiphora myrrha (Nees) Engl. gum-resin on hepatorenal injury in diabetic rats.

Materials and methods

Chemical and reagents

Streptozotocin (STZ) (purity ≥ 98 %) was purchased from Sigma Aldrich Company (St Louis, MO, USA). Metformin, hematoxylin, and Eosin were procured from Merck Company (Darmstadt, Germany). Ketamine hydrochloride was purchased from Laboratories Strop NV (Brussels, Belgium), and xylazine was bought from Interchemie Werken de Adelaar Company (Venray, Netherlands). A periodic acid Schiff kit was obtained from Asiapajohesh Company (Amol, Iran), and a rat insulin kit was purchased from Mercodia Company (Uppsala, Sweden). Dimethyl sulfoxide (DMSO) (purity ≥ 99.9 % ) was obtained from Carlo ERBA Company (Val de Reuil, France). The other used kits for biochemical assessments were obtained from Bionik Company (Tehran, Iran).

Extract preparation

Myrrh was procured from a traditional plants market in Birjand, Iran. A sample of purchased myrrh was tested (nitric acid and carbon disulfide color reaction) and identified as a myrrh resin, and a sample was stored (voucher number: 527) in the herbarium of the University of Birjand, Iran.

The gum-resins were powdered with an electric miller. Extraction was performed using a conventional maceration method with continuous stirring at room temperature for 48-h. In brief, 100 g of the powdered myrrh was macerated in 900 mL of 80 % ethanol (1:10 w/v) and then filtered using filter papers (Whatman No. 4, England). The filtrate was concentrated to one-sixth of the original volume by a rotatory vacuum evaporator (Wiggens, Italy). The resulting solution was pre-frozen (-60 °C) for 8 h prior to freeze-drying in a lyophilization apparatus (Dena Vacuum Industry, model FD-5005-BT, Iran) [12]. The extraction yield was 9.5 %. Freshly before use, the extract was dissolved in a 0.9 % saline solution containing 0.1 % DMSO.

Animals

Healthy adult male Wistar rats (8-week old) were purchased from the animal house of Birjand University of Medical Sciences, Iran. They were kept in a temperature (24 ± 2 ºC) and humidity (30–35 %) -controlled environment with a 12-h light/dark cycle. All animals had ad-libitum access to food (Behparvar Co, Iran) and tap water during the study period.

Diabetes induction and experimental design

In order to induce diabetes, rats were fasted overnight with free access to water. Then, STZ was freshly dissolved in clod citrate buffer (0.1 M, pH 4.5) and injected intraperitoneally (i.p.) to the fasted rats at a single dose of 55 mg/kg [20]. Meanwhile, control animals (n = 8) received 0.5 mL of 0.1 M citrate buffer (i.p.) as vehicle. Diabetes induction was confirmed by determining fasting blood glucose (FBG) three days post-STZ injection. Animals with FBG levels equivalent to or greater than 300 mg/dL were considered diabetic [21]. In order to ensure the establishment of hepatic and renal damage in diabetic rats, they were kept untreated for 4-week [22].

A total of 48 rats (40 diabetic and 8 non-diabetics) were randomly distributed into six groups (n = 8):

  • Group 1: Non-diabetic control rats (NC).

  • Group 2: STZ-induced diabetic model (DM).

  • Group 3: Diabetic rats treated with metformin at the dose of 500 mg/kg (MET500).

  • Group 4: Diabetic rats treated with EEM at the dose of 100 mg/kg (EEM100).

  • Group 5: Diabetic rats treated with EEM at the dose of 300 mg/kg (EEM300).

  • Group 6: Diabetic rats treated with EEM at the dose of 500 mg/kg (EEM500).

NC and DM groups were treated with vehicle solution (normal saline with 0.1 % DMSO). All treatments were performed orally, once per day, for 28 days. According to previous studies, the metformin dose was selected [23]. The doses of EEM were carefully chosen based on previous studies [18, 19, 24, 25].

Sample collection

On the 27th day of study, 24-hour urine samples of all animals were collected by placing them into metabolic cages. The next day, following 8-h fasting, rats were anesthetized with ketamine-xylazine injection (65:10 mg/kg, i.p.), their hearts were punctured with an injection needle’s tip, and whole blood samples were collected [26]. Immediately after blood collection, the left kidney and liver were dissected out, weighed, and fixed in 4 % paraformaldehyde solution. Prior to carcasses disposal, all rats were decapitated in order to ensure death.

Biochemical assay

Plasma levels of the FBG, urea, creatinine (Cr), 24-hour urine protein (UTP), aspartate transaminase (AST), and alanine transaminase (ALT) were assessed using standard diagnostic kits (Bionik, Iran) adapted for a Roche Hitachi 912 auto-analyzer machine (Japan). Furthermore, the plasma insulin concentration was measured by a commercial ELISA kit (Mercodia) according to the manufacturer’s instructions.

Histopathological evaluations

The fixed samples were embedded in paraffin wax and sectioned at 5 μm for hematoxylin-eosin and periodic acid-Schiff (PAS) staining methods. Ten sections of liver/kidney were selected randomly from each group and evaluated under a light microscope (Euromex-CMEX-10, Netherlands). Pathological lesions (hemorrhage, congestion, infiltration, and degeneration) were evaluated using a scoring checklist (0 = none, 1 = mild, 2 = moderate, 3 = severe) in a blinded manner [27–29].

Statistical analysis

All values are presented as mean ± standard deviation. Data were analyzed using the statistical software IBM SPSS version 22. The normality of data was checked using the Shapiro-Wilk normality test. The differences between groups were determined with ANOVA and Dunnett’s T3 post hoc tests. The Kruskal-Wallis test was used to compare the pathological scores between the studied groups. Differences were inferred as significant when the P-value was < 0.05.

Results

Effects on body weight and organ weights

In the DM group, body weight (B.wt) was decreased (p < 0.001, Table 1); whereas, liver index (liver weight to B.wt) and kidney index (kidney weight to B.wt) were increased (p < 0.001 and p < 0.05 respectively) compared to the NC group. Treatment with metformin (MET500) and EEM (all doses) could not affect B.wt and the elevated liver index. The kidney indexes were significantly lower in the EEM100 (p = 0.033) and MET500 (p = 0.015) groups than DM group.

Table 1.

Effects of ethanolic extract of myrrh on body weight, liver and kidney weights in diabetic rats

Groups Body weight (g) Liver to body weight ratio (*100) Kidney to body weight ratio (*1000)
NC 263.33 ± 34.33a 3.00 ± 0.40a 3.29 ± 0.31a
DM 167.83 ± 36.86b 5.94 ± 0.85b 7.94 ± 1.98b
MET500 178.00 ± 29.69 b 4.67 ± 1.10 b 5.78 ± 1.29c
EEM100 174.16 ± 17.57 b 5.01 ± 1.06 b 5.57 ± 1.05c
EEM300 159.40 ± 18.10 b 5.21 ± 0.66 b 6.07 ± 0.99b
EEM500 178.33 ± 29.15 b 5.43 ± 0.35 b 6.00 ± 0.37b
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Values are expressed as mean ± SD (n = 8). Differences among groups were analyzed by a one-way analysis of variance (ANOVA) followed by Tukey’s post hoc test. Different letters (a-c) show a significant difference (p < 0.05) between groups. NC: normal control group; DM: STZ-induced diabetic model group; MET500: diabetic rats treated with metformin at dose of 500 mg/kg; EEM100-EEM500: diabetic rats treated with 100–300 mg/kg ethanolic extract of myrrh respectively

Effects on FBG, insulin, and liver enzymes

Table 2 shows the results of FBG, plasma insulin, AST, and ALT in different studied groups. Diabetic rats exhibited higher FBG and lower insulin levels than the NC group (p < 0.001, both). No significant differences in FBG and insulin levels were found between EEM-treated rats and the DM group. Compared to the DM group, treatment with metformin significantly decreased FBG levels (p < 0.001) and enhanced the plasma insulin concentration (p = 0.002) in diabetic rats. Assessment of liver enzymes revealed that AST and ALT levels were higher in the DM group than the NC group (p < 0.001, each). Neither metformin nor EEM could reduce the elevated AST levels. Unlike EEM (all doses), metformin treatment attenuated ALT levels in diabetic rats.

Table 2.

Effects of ethanolic extract of myrrh on fasting blood glucose, plasma insulin, and liver enzymes concentrations in diabetic rats

Groups Glucose (mg/dL) Insulin (ng/uL) AST (IU/L) ALT (IU/L)
NC 98.33 ± 8.84 a 2.55 ± 0.44a 49.50 ± 4.16 a 29.33 ± 8.82 a
DM 447.33 ± 74.03 b 0.68 ± 0.12 b 85.16 ± 11.14 b 62.33 ± 8.21 b
MET500 163.16 ± 21.59 c 1.29 ± 0.08 c 72.33 ± 10.38 b 31.66 ± 21.43 a
EEM100 437.00 ± 50.34 b 0.44 ± 0.20 b 67.66 ± 21.34 b 64.16 ± 19.83 b
EEM300 457.50 ± 61.86 b 0.59 ± 0.09 b 67.60 ± 18.46 b 57.80 ± 18.39 b
EEM500 486.33 ± 42.08 b 0.78 ± 0.16 b 80.66 ± 13.58 b 70.60 ± 8.01 b
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Values are expressed as mean ± SD (n = 8). Differences among groups were analyzed by a one-way analysis of variance (ANOVA) followed by Tukey’s post hoc test. Different letters (a-c) show a significant difference (p < 0.05) between groups. NC: normal control group; DM: STZ-induced diabetic model group; MET500: diabetic rats treated with metformin at dose of 500 mg/kg; EEM100-EEM500: diabetic rats treated with 100–300 mg/kg ethanolic extract of myrrh respectively; AST: aspartate aminotransferase; ALT: alanine aminotransferase

Effects on Cr, urea, urine volume, and UTP

The results of renal function tests, including plasma Cr and urea, 24-h urine volume, and UTP, are presented in Table 3. Diabetes caused significant increases in plasma Cr, urea, urine volume, and 24-h UTP compared to the NC group (p < 0.001, all). Plasma Cr only restored in metformin-treated animals. Treatment with EEM at all doses could not ameliorate Cr in diabetic rats. Neither Metformin nor EEM (at all doses) could mitigate blood urea in diabetic rats. The 24-h urine volume significantly decreased in MET500, EEM300, and EEM500 groups compared to the DM group. Treatment with EEM at the dose of 500 mg/kg could decrease urine volume close to its level in the NC group. Compared to the NC group, DM (p < 0.001), EEM300 (p = 0.02), and EEM500 (p < 0.001) had significantly higher UTP exertion. Only metformin and EEM at the dose of 100 mg/kg had significantly lower UTP than the DM group.

Table 3.

Effects of ethanolic extract of myrrh on renal function tests and 24-hour urine volume in diabetic rats

Groups Cr (mg/dL) Urea (mg/dL) 24-h urine volume (mL) UTP (mg/24-h)
NC 0.95 ± 0.05a 33.66 ± 3.82a 7.83 ± 1.68 a 9.00 ± 1.41 a
DM 1.08 ± 0.09b 71.50 ± 16.83 b 22.83 ± 3.72 b 21.00 ± 5.21b
MET500 0.90 ± 0.08a 80.33 ± 7.58 b 11.38 ± 4.74c 12.33 ± 4.03a
EEM100 1.13 ± 0.05b 63.33 ± 5.95 b 20.69 ± 3.07 b 11.83 ± 4.26a
EEM300 1.12 ± 0.08b 86.66 ± 13.07 b 15.33 ± 2.63d 16.66 ± 3.50 b
EEM500 1.14 ± 0.05b 73.83 ± 8.61 b 8.38 ± 2.63 a 22.83 ± 3.76 b
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Values are expressed as mean ± SD (n = 8). Differences among groups were analyzed by a one-way analysis of variance (ANOVA) followed by Tukey’s post hoc test. Different letters (a-d) show a significant difference (p < 0.05) between groups. NC: normal control group; DM: STZ-induced diabetic model group; MET500: diabetic rats treated with metformin at dose of 500 mg/kg; EEM100-EEM500: diabetic rats treated with 100–300 mg/kg ethanolic extract of myrrh respectively; Cr: cratinine; UTP: urine total protein

Effects on liver histopathology

The liver histological structures of specimens belong to the NC group showed normal morphology. In the NC group, separate hepatocytes were radially organized over central veins, and the portal triads bordered each hepatic lobule. Besides, sinusoidal spaces without hemorrhage and dilatation were observed in the NC group (Fig. 1A). Conversely, the DM group showed severe morphological changes. These alterations included some degree of fatty changes (microvesicular fatty degenerations, ballooning of hepatocytes) and mononuclear cell infiltration (Fig. 1A). Semi-quantitative assessments revealed that in the DM group, the mean score of liver histological changes, including congestion (p < 0.001), hemorrhage (p < 0.01), infiltration (p < 0.01), and degeneration (p < 0.001) were significantly increased compared to the NC group. Neither metformin (500 mg/kg) nor EEM (100–500 mg/kg) could effectively ameliorate all the elevated pathological scores except for degeneration (Fig. 1B). EEM treatment only at the dose of 500 mg/kg showed a mild degree of efficacy by reducing the score of degenerative alterations (mainly fatty changes); however, some degrees of pathological lesions such as sinusoidal hemorrhage and inflammatory cells infiltration were still detectable in the EEM500 group.

Fig. 1.

Fig. 1

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Histological examination of liver sections of normal control (NC), diabetic model (DM), and diabetic rats treated with 500 mg/kg metformin (MET500), and diabetic groups treated with ethanolic extract of myrrha gum at the doses of 100–500 mg/kg (EEM100- 500). (A) Photomicrographs of liver sections stained with Hematoxylin and eosin dyes (400 X, scale-bars = 50 μm). CV: central vein, red arrow marks a hepatocyte, yellow arrow indicates a sinusoid, green arrowheads indicate mononuclear cells infiltration, brown arrows show hepatocytes with early fatty changes, and white arrows indicate sinusoidal hemorrhage. (B) Boxplots show histological scores of liver congestion, hemorrhage, infiltration and degeneration. Scoring was done as none (0), low (1), mild (2) and severe (3).*p < 0.01 vs. normal control group, # p < 0.05 vs. diabetic group

Effects on Kidney histopathology

The histological appearance of PAS-stained kidney sections was normal in the NC group. Histological evaluation of kidney sections of the DM group showed severe morphological degenerations. The prominent observed renal alterations were tubular defects, mononuclear cells infiltration, mesangial matrix expansion, and glomerular hyalinosis (Fig. 2A). Significant increases in histopathological scores of congestion (p < 0.05), hemorrhage (p < 0.05), infiltration (p < 0.01), and degeneration (p < 0.01) were detected in the DM group compared to the NC group (Fig. 2B). Metformin significantly eliminated the score of degenerative alterations in diabetic rats (p = 0.03). EEM (300–500 mg/kg) could slightly attenuate kidney congestion score in a dose-dependent manner, but had no remarkable effect on the other histopathological alterations.

Fig. 2.

Fig. 2

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Histological examination of kidney sections of normal control (NC), diabetic model (DM), and diabetic rats treated with 500 mg/kg metformin (MET500), and ethanolic extract of myrrha gum at the doses of 100–500 mg/kg (EEM100- 500). (A) Photomicrographs of kidney sections stained with periodic acid Schiff technique (400 X, scale-bars = 50 μm). Red arrow marks a glomerulus, red arrowhead indicates Bowman’s capsule, green arrows mark proximal convoluted tubules and black arrow indicates a distal convoluted tubule, yellow arrows show glomerular hyalinosis, yellow arrowheads indicate mesangial matrix expansion, and green stars mark tubular defects mostly vacuolated tubular epithelium. (B) Boxplots show histological scores of kidney congestion, hemorrhage, infiltration and degeneration. Scoring was done as none (0), low (1), mild (2) and severe (3).* p < 0.01 vs. normal control group, # p < 0.05 vs. diabetic group

Discussion

Diabetic nephropathy and hepatopathy were well established in diabetic rats, as indicated by altered biochemical and histological parameters. Diabetic animals had significantly higher plasma AST, ALT, urea, Cr, urine volume, and UTP than the control rats. The histopathological findings revealed that liver and kidney architectures were extensively altered in diabetic animals.

Our findings demonstrated that EEM could not influence glucose metabolism and insulin production in diabetic rats. Contrary to these findings, Sotoudeh and coworkers have reported that Commiphora myrrha extract (438 mg/kg) decreased blood glucose levels in diabetic rats [18]. In the mentioned study, an herbal combination formula consisted of Commiphora mukul, Commiphora myrrha, and Terminalia chebula has been used. Hence, the possible reason for this inconsistency could be attributed to the hypoglycemic activity/s of other used herbs or their interaction. These findings are in agreement with our previous study in which the EEM could not decrease FBG levels in diabetic rats [19].

Diabetic animals also showed elevated relative liver and kidney weights. Increasing the liver weight could be caused by increased triglyceride deposition in the liver due to hypoinsulinemia. Moreover, kidney index elevation might be attributed to several factors such as overexpression of epidermal growth factor (EGF), transforming growth factor-beta (TGF-β1), and vascular endothelial growth factor (VEGF) [10, 11, 28–30]. Contrary to our findings, it has been previously reported that 14 weeks of oral administration of myrrh extract (500 mg/kg) could decrease elevated liver weight in high-fat diet treated rats [25]. Moreover, it has also been reported that 2-week oral administration of myrrh extract (500 and 1000 mg/kg) could reduce liver and kidney relative weights in diabetic rats [31]. A possible explanation for this discrepancy may be the differences in the investigation time and study design. Orabi et al. have investigated myrrh extract for 14 weeks; however, our treatment lasted for 28 days in the present study. AL-Yahya and coworkers have started their experiment 72-h post diabetes induction and lasted it for two-week. Evidence demonstrates that at least four weeks is needed to develop diabetic complications such as diabetic nephropathy in rats [10, 22].

Diabetic animals had significantly increased liver enzymes levels (AST and ALT) than control rats, which are in good agreement with previous reports [11, 12, 32]. Liver enzymes are normally presented in the cytosol and due to hepatocyte plasma membrane damage leaked out into the bloodstream [27, 33]. EEM treatment could not ameliorate the elevated levels of these enzymes. A previous study has shown that 60 days administration of ethanolic extract of Commiphora mukul gum-resin (200 mg/kg), a similar species to C. myrrha, significantly ameliorated tissue concentrations of AST and ALT [34]. There are several possible explanations for these results. Rats with FBG > 126 mg/dL have been used by Ramesh et al., while in our experiment, diabetic rats with FBG levels above 350 mg/dL were investigated. It is well demonstrated that in a rat model of STZ-induced diabetes model, the FBG levels reflect the severity of associated complications. In this model, FBG concentrations are categorized as moderate (120 and 300 mg/dL), which is often not severe enough for development of diabetes-related complications, and severe (FBG > 300 mg/dL) [21]. Besides, in the mentioned study, the extract has been investigated for 60 days. However, in the present study, our treatment lasted for 28 days. They also assessed tissue levels of AST and ALT, whereas we evaluated plasma levels of liver enzymes.

In line with biochemical parameters, liver pathology of diabetic rats revealed several histological alterations such as microvesicular fat vacuoles deposition (fatty change), hemorrhage, and mononuclear cell infiltration. The main underlying mechanisms of diabetes that contribute to liver damage are increased oxidative stress and aberrant inflammatory response [12]. Liver tissues of diabetic rats treated with EEM also showed several pathological lesions. However, the severity of alterations was slightly lower than in untreated diabetic rats. EEM treatment (500 mg/kg) significantly ameliorated the degenerative changes (fatty change) score, but other lesions such as sinusoidal hemorrhage and inflammatory cell infiltration were still detectable. Similarly, Orabi and coworkers have reported that 14-week simultaneous administration of EEM at a dose of 500 mg/kg with a high-fat diet slightly prevented fatty change diffusion and vessels congestion in hepatic tissues of rats [25].

Diabetic animals had significantly higher urea, Cr, urine volume, and UTP excretion than the control rats. These findings are in good agreement with previous reports showing that diabetic nephropathy was well established [10, 11]. EEM treatment could not ameliorate the elevated plasma urea and Cr levels. However, it significantly decreased urine volume in a dose-dependent manner. EEM treatment only at the dose of 100 mg/kg could significantly decrease UTP excretion. Similarly, it has been reported two weeks of treatment with EEM at doses of 200, 400, and 600 mg/kg significantly increased plasma Cr in male Wistar rats [35]. In addition, it has been reported that myrrh ingestion (1000–5000 mg/kg) could increase Cr levels in male Nubian goat kids [36]. To the best of our knowledge, no study has been yet investigated the effect of myrrh on urine volume and UTP. However, there is evidence showing that myrrh has been used as an anti-diuretic agent for the treatment of nocturnal enuresis [37].

Histological study of kidneys revealed severe glomerular and tubular defects in diabetic rats. Despite the slight positive effect of EEM in preventing tubular congestion, it could not ameliorate other pathological lesions such as mesangial matrix expansion, mononuclear cell infiltration, and hyalinosis. To our knowledge, no study has been yet performed to study the myrrh effect of kidney pathology in diabetic conditions. However, there is a study in which the protective efficacy of myrrh has been reported against cadmium chloride and methotrexate-induced renal damage in rats [38, 39].

It worth be noted that the plant Commiphora myrrha (Nees) Engl. does not grow in Iran. In consequence, we could not directly collect the myrrh and control important parameters influencing its quality, such as time of collection and storage condition. On the other hand, due to its importance in Traditional Persian Medicine, several gummy substances that resemble myrrh are available in the traditional markets [40]. In the present study, myrrh was obtained from a highly reputed traditional market and was preliminarily evaluated by colorimetric tests. However, we are unaware of its source, harvested area, and probable contamination with pesticides or heavy metals. Therefore, the difference between our results with previous literature might be attributed to the quality of the myrrh sample used in this study.

Curzerene, beta selinene, germacrene B, limonene, isocericenine, myrcenol, and spathulenol are the main phytochemical constituents identified in the myrrh [41]. In a study conducted by Hosseinkhani and coworkers, the chemical constitution of six myrrh samples from Iran (n = 3) and the United Arab Emirates (n = 3) traditional markets has been investigated. They reported furanoeudesma-1, 3-diene, curzerene, and lindestrene as the main constituents of the myrrh essential oil, which was in line with previous studies [42, 43]. They could not found significant differences in myrrh essential oil ingredients between the samples [40]. Therefore, myrrh’s most profound organic constituents (both in essential oil and ethanolic extract) are sesquiterpenes such as curzerene, furanodiene, and lindestrene. The compounds are mostly known for their antibacterial, antifungal, local anesthetic properties [44].

Although in the present study, hepatorenal effects of EEM were investigated in a well-developed animal model of diabetes, this study still suffers from some limitations. Since myrrh is used in several traditional medicine formulas, it would have been better to investigate its combination efficacy with metformin.

Conclusions

The main conclusion drawn from the present study’s findings is that EEM could not attenuate diabetes-associated pathological characteristics of liver and kidney in rats.

Acknowledgements

We thank Dr. Farrokhfall for helping us in insulin assay test.

Abbreviations

Myrrh

Commiphora myrrha oleo-gum

EEM

Ethanolic extract of myrrh

AST

Aspartate transaminase

ALT

Alanine transaminase

UTP

Urine total protein

STZ

Streptozotocin

i.p.

Intraperitoneal.

FBG

fasting blood flucose

NC

normal control

DMSO

Dimethylsulfoxide

MET

Metformin

DM

Diabetic model

Cr

Creatinine

PAS

Periodic acid-Schiff

B.wt

Body weight

TGF-β1

Transforming growth factor-Beta1

EGF

Epidermal growth factor

VEGF

Vascular endothelial growth factor

Authors’ contributions

M.HT and M.H conceived the idea of research and designed the study. M.S performed the animal experiments. K.V-D and M.M performed biochemical and pathological tests. M.H analyzed the data and wrote the manuscript draft. All authors read and approved the final manuscript.

Funding

This work was partially supported by Birjand University of Medical Sciences (Grants No.:455159).

Data availability

The datasets used and/or analyzed during the current study are available from the corresponding author on reasonable request.

Declarations

Ethics approval and consent to participate

All study protocols were approved by the Birjand University of Medical Sciences Ethics Committee (permit code: Ir.bums.REC.1396.16).

Consent for publication

Not applicable.

Conflict of interest

All authors declare that they have no competing interest.

Footnotes

Publisher’s note

Springer Nature remains neutral with regard to jurisdictional claims in published maps and institutional affiliations.

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Associated Data

This section collects any data citations, data availability statements, or supplementary materials included in this article.

Data Availability Statement

The datasets used and/or analyzed during the current study are available from the corresponding author on reasonable request.

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