Quercetin potentiates transdifferentiation of bone marrow mesenchymal stem cells into the beta cells in vitro.

PURPOSE: Type 1 diabetes is an autoimmune disease caused by the destruction of ß-cells in the pancreas. Bone marrow mesenchymal stem cells are multipotent and easy accessible adult stem cells that may provide options in the treatment of type 1 diabetes. Injured pancreatic extract can promote the differentiation of rat bone marrow mesenchymal stem cells into ß-cells. We aimed to observe the effect of quercetin in differentiation and insulin secretion in ß-cells. METHODS: Bone marrow mesenchymal stem cells were obtained from the tibiae of rats. Cell surface markers were analyzed by flow cytometry. The cells were treated with rat injured pancreatic extract and quercetin for 2 weeks. Insulin secretion was measured by ELISA. Insulin expression and some islet factors were evaluated by RT-PCR. PDX1, a marker for ß-cell function and differentiation, was evaluated by both immunocytochemistry and Western blot. ß-cell count was determined by stereology and cell count assay. RESULTS: ELISA showed significant differences in insulin secretion in the cells treated with RIPE + 20 µM quercetin (0.55 ± 0.01 µg/L) compared with the cells treated with RIPE alone (0.48 ± 0.01 µg/L) (P = 0.026). RT-PCR results confirmed insulin expression in both groups. PDX1 protein was detected in both groups by Western blot and immunocytochemistry. Stereology results showed a significant increase in ß-cell number in the RIPE + quercetin-treated cells (47 ± 2.0) when compared with RIPE treatment alone (44 ± 2.5) (P = 0.015). CONCLUSIONS: Quercetin has a strengthening effect on the differentiation of rat bone marrow mesenchymal stem cells into ß-cells and increases insulin secretion from the differentiated ß-cells in vitro.

EFFECTS OF ARNEBIA EUCHROMA EXTRACT ON STREPTOZOTOCIN INDUCED DIABETES IN RATS: A STEREOLOGICAL STUDY.

BACKGROUND: Diabetes mellitus (DM) is considered as an important health confounder in our world, which necessitates its better management by new methods. In this study, we have evaluated the effects of oral Arnebia Euchroma (AE) extract on different stereological parameters of the pancreas as well as blood glucose in Streptozotocin (STZ)-induced diabetes in rats. METHODS: We divided 48 Wistar rats into 4 groups: C1 including normal rats, C2 not-treated diabetic rats, E1 with diabetic rats receiving 100 mg/kg AE extract orally, and E2 including diabetic rats treated with 300 mg/kg AE extract. Stereological study was done and the levels of blood glucose were also estimated and compared between experimental and control groups. RESULTS: There were significant differences in volumes of pancreatic islets, ß cell populations, blood glucose levels in AE treated groups compared with not-treated diabetic group. CONCLUSION: Although AE did not completely prevent or heal the pancreatic damage, its oral administration showed promising effects on maintaining the population of beta cells, the main insulin secreting cells, after STZ-induced injury and also lowered blood glucose levels compared to the not-treated diabetic group.

Stereological study of the effect of black olive hydroalcoholic extract on osteoporosis in vertebra and tibia in ovariectomized rats.

UNLABELLED: A cocktail of many different antioxidants might be more effective than supplementation with a single molecule, and it closely resembles the natural environment in which active compounds were found. This is the first study well-grounded in stereological examination that showed that black olive extract effectively can ameliorate the quantitative changes of the bone structure and prevented bone loss in this osteoporosis animal model. INTRODUCTION: The aim of this study was to quantitatively evaluate the effects of black olive extract consumption on treatment of ovariectomized (OVX) induced osteoporosis in rats. This is the first study well-grounded in stereological examination. METHODS: Ninety adult rats were allocated to control, sham-operated, OVX, and olive-supplemented OVX groups (received 250-, 500-, and 750-mg/kg body weight black olive hydroalcoholic extract orally) for 16 weeks. At the end of the experiment, blood samples were collected, and plasma levels of calcium, phosphorus, and alkaline phosphatase were assayed. Then, the specimens from both the tibia and fifth lumbar vertebra (L5) bones were processed, and stereological analysis was performed. RESULTS: Administration of extract resulted in decrease of alkaline phosphatase level during the treatment. After treatment of OVX rats with three doses of extract, the total number of the osteocytes revealed an increment in 500- and 750-mg/kg treated groups in comparison to the OVX group. This increment was significant only in L5. Compared to the OVX group, a significant increase was observed in the number of osteoblastsin L5 vertebra in three doses of extract-treated groups. However, this increment in tibia was statistically significant only in 750-mg/kg black olive hydroalcoholic extract-treated group. Moreover, the number of osteoclast cells were significantly decreased in vertebra and tibia in the treated groups compared to the OVX group (P < 0.05). CONCLUSION: Black olive hydroalcoholic extract effectively can ameliorate the quantitative changes of the bone structure and prevented bone loss in this osteoporosis animal model. Thus, it can be a promising candidate for treatment of accelerated bone loss especially in postmenopausal osteoporosis.

Sodium metabisulphite, a preservative agent, decreases the heart capillary volume and length, and curcumin, the main component of Curcuma longa, cannot protect it.

Sodium metabisulphite is used as an antioxidant agent in many pharmaceutical formulations. It is extensively used as a food preservative and disinfectant. It has been demonstrated that sulphite exposure can affect some organs. Curcumin, the main element of Curcuma longa, has been identified to have multiple protective properties. The present study extends the earlier works to quantitative evaluation of the effects of sulphite and curcumin on the heart structure using stereological methods. In this study, 28 rats were randomly divided into four experimental groups. The rats in groups I to IV received distilled water (group I), sodium metabisulphite (25 mg/ kg/day) (group II), curcumin (100 mg/kg/day) (group III), and sodium metabisulphite+curcumin (group IV), respectively, for 8 weeks. The left ventricle was subjected to stereological methods to estimate the quantitative parameters of the myocardium. A 20 % decrease was observed in the total volume of ventricular tissue in the sulphite-treated animals compared to the distilled water treatment (P < 0.02). Also, the volume and length of the capillaries were reduced by 43 % on average in the sulphite-treated rats in comparison to the distilled water-treated animals (P < 0.02). However, no significant change was seen in the mean and total volume of the myocardium and the cavity and diameter of the capillaries after sulphite ingestion. Treatment with curcumin did not protect the animals against the structural changes of the ventricle. Sulphite, as a preservative food agent, reduced the length and volume of the ventricular capillaries and curcumin could not protect them.

Effect of intradermal injection of heparinase III on skin wound healing in diabetic rats.

AIM: neuropathy and vascular damage in this disease. Heparanase is an endoglycosidase that degrades heparan sulfate in the extracellular matrix and is believed to promote angiogenesis. The present study has been performed to investigate the effect of heparinase III (an enzyme which exclusively cleaves heparan sulfate) on wound healing in diabetic rats. METHODS: The rats became diabetic by a single streptozotocin injection. Two weeks later, a wound was created by excision of the skin in the left paravertebral area. Heparinase III (0.2 unit) was injected intradermally around the wound every 5 days, starting on day one, for a total of three doses. The wound area was measured every 3 days. After completion of wound healing, full thickness skin samples were taken from the wound sites and evaluated for volume density of the collagen bundles, numerical density of the fibroblasts, and length density of the vessels. RESULTS: Heparinase III accelerated wound closure compared to control diabetic animals. Microscopical examination revealed that it increased angiogenesis with no significant effect on collagen density and the number of fibroblasts. CONCLUSION: Heparinase III induces angiogenesis and improves wound healing in diabetic animal model.

Ameliorative effects of curcumin on the spermatozoon tail length, count, motility and testosterone serum level in metronidazole-treated mice.

Metronidazole (MTZ) is used as an antiparasitic drug. Curcumin is considered as anti-oxidant and anti-inflammatory agent. The ameliorative effects of curcumin on MTZ induced toxicity on mice spermatozoon tail length, count, motility and testosterone level were investigated. MTZ was administered in 500 and 165 (high and therapeutic doses) mg/kg/day, with and without curcumin (100 mg/kg/day). After 16 days the above parameters were assessed. Spermatozoon count and motility and serum testosterone level MTZ-treated (500 and 165) mice were reduced. In the mice treated with MTZ+curcumin these parameters decreased but in a lesser extent than the MTZ-treated animals. Mid-piece and total lengths of the spermatozoon tail in control animals were 31.6 ± 9.0 µm and 100.3 ± 15.0 µm and in the mice treated with high doses (500) of MTZ were reduced. The mid-piece and total spermatozoon tail length has been decreased in a lesser extent in the mice treated with high dose MTZ+curcumin than the mice treated with high dose MTZ (p<0.01). But the length was not changed in animals treated with therapeutic dose of MTZ. It means curcumin treated animals had ~52% and ~39% average increase in mid-piece and total lengths in comparison with the MTZ-treated (500) animals. Stereological estimation of the sperm tail length, including sampling of spermatozoa and also counting of the intersections of their tails with the stereological grids was a rapid technique and took only 5-10 minutes. It can be concluded that curcumin has an ameliorative effect on the spermatozoon, testosterone level and tail length in MTZ-treated mice.

Sertraline and curcumin prevent stress-induced morphological changes of dendrites and neurons in the medial prefrontal cortex of rats.

Stress induces structural and behavioral impairments. The changes in dendrites and neurons are accompanied by impairments in the tasks mediated by the medial prefrontal cortex (mPFC). The present study was conducted to evaluate the structural changes of the dendrites and neurons of the mPFC after stress using stereological methods. In addition, the effects of a natural and a synthetic substance, i.e., curcumin and sertraline, were evaluated. The rats were divided into 7 groups: stress + distilled water, stress + olive oil, curcumin (100 mg/kg/day), sertraline (10 mg/kg/day), stress + curcumin, stress + sertraline, and control groups. The animals were submitted to chronic variable stress for 56 days. The results showed an average 15% reduction in the length of the dendrites per neuron in the mPFC after stress (p < 0.004). The total spine density was reduced by 50% in the stress (+ olive oil or + distilled water) groups in comparison with the control group (p < 0.01). The main reduction was seen in the thin and mushroom spines, while the stubby spines remained unchanged. Mean volume and surface area of the neurons were decreased by 14% and 10% on average in the stress (+ distilled water or + olive oil) rats in comparison to the control rats, respectively (p < 0.01). The data revealed that treatment of stressed rats with curcumin or sertraline can prevent the loss of spines and reduction of dendrite length, volume and surface area of the neurons. Sertraline and curcumin can prevent structural changes of the neurons and dendrites induced by stress in the mPFC of rats.

Effects of low-intensity pulsed ultrasound on healing of mandibular bone defects: an experimental study in rabbits.

Research evidence suggests that low-intensity pulsed ultrasound (LIPU) produces significant osteoinductive effects, accelerating the healing of bone defects. The authors investigated the effects of LIPU on mandibular bone defects in a rabbit model. Fifty-six adult Dutch rabbits were divided randomly into control, LIPU-1 (1MHz), and LIPU-3 (3MHz) groups. A mandibular defect was created in all rabbits. The effect of LIPU on mandibular defects was assessed by frequency (1 or 3MHz) and timing (2 and 4 weeks). Bone mineral density (BMD) was measured and stereology and histology performed; results were compared at the end of 2 and 4 weeks. LIPU-3 resulted in significantly higher bone formation compared to the control group at the end of week 4 on histological assessment (P=0.008). BMD was significantly higher at 4 weeks than at 2 weeks (P=0.03). LIPU-3 increased the numerical density of osteoblasts and osteocytes at the end of week 4 (P=0.05 and P=0.001, respectively). The results of this study are in favour of using LIPU 3MHz to accelerate mandibular bone healing. However, this study suggests that a frequency of 3MHz and the longer application of LIPU 3MHz for 4 weeks can only promote 8% mandibular bone healing in rabbits. Therefore, the use of LIPU has no really convincing, consistent clinical effects on maxillofacial bone healing.

Stereological study on the submandibular gland in hypothyroid rats.

Most research done on hypothyroidism has focused on physiological and biochemical aspects of the tissues, whilst there has been little work on tissue morphology especially on salivary glands. The present study has used hypothyroid Wistar rats as a model for investigating the effects of hypothyroidism on submandibular gland structure. Two groups of Wistar rats were studied. One was made hypothyroid with methimazole and the second was an untreated euthyroid group (control). They were euthanised after 10 weeks. Submandibular glands were removed and studied. Systematic random transverse sections were obtained from submandibular glands and subjected to morphometric analysis. Volume density and absolute volume of granular, striated and excretory ducts and interlobular connective tissue were estimated by point counting. Volume-weighted mean particle volume of serous and mucous acini was also determined by unbiased stereology. Absolute volume of granular ducts in the submandibular gland of hypothyroid rats was reduced by approximately 50% (p<0.009) whilst that of the striated and excretory ducts and interlobular connective tissue was unaffected. Volume of serous acini was also significantly (p<0.03) lower in hypothyroid rats. These changes suggest that hypothyroidism has an effect on submandibular gland structure, and that this effect occurs mostly in two major exocrine compartments (granular duct and serous acinus) of the gland.

Stereological study of the cells of dorsal root ganglia in male diabetic rats.

Most research on diabetes mellitus has focused on physiological and biochemical aspects of the peripheral nervous system, whilst little work has been done on morphological changes of the neurons. In the present study the effects of diabetes mellitus on cervical and lumbar dorsal root ganglia (C7 and L5) were investigated using modern stereological methods. Twelve adult male Sprague-Dawley rats were randomly divided into two groups. Each group contained six male rats. Diabetes was induced in the experimental group by intraperitoneal injection of 50 mg/kg streptozotocin. At the end of 6 weeks, the rats were fixed by whole body perfusion transcardially with a buffered formalin solution. The seventh cervical and fifth lumbar dorsal root ganglia were removed and immersed in buffered formalin. After tissue processing, the ganglia were embedded in cylindrical paraffin blocks. Isotropic uniform random sections were obtained using the orientator method. Sections (5 microm thick) were selected and stained with Heidenhain's azan. Volume of perikarya of A- and B-cells and their nuclei was estimated using the nucleator method. Before estimating the mean volume, the cells were sampled using the physical disector and point sampling method. Measurements showed that mean perikaryal and nuclear volume of A- and B-cells of dorsal root ganglia (C7 and L5) was reduced in diabetic rats (p<0.05). B-cell mean perikaryal volume in diabetic rats and A- and B-cell mean nuclear volume were reduced by 66% on average. The mean volume of A-cell perikarya was affected less than the others (average 33%). In addition, the difference between the perikaryal and nuclear volume of the seventh cervical and fifth lumbar dorsal root ganglia was not statistically significant. The present study, using stereological techniques, demonstrates reduced perikaryal and nuclear volume of the seventh cervical and fifth lumbar dorsal root ganglia in diabetic rats.

The healing effect of arnebia euchroma in second degree burn wounds in rat as an animal model.

BACKGROUND: Finding more efficient agents with fewer side effects for treatment of burns has always been a concern for researchers. Silver sulfadiazine (SSD), apparently due to its antimicrobial effect, is still one of the most common prescribed agents. Previous studies suggested that Arnebia euchroma (AE) has shown antimicrobial and anti-inflammatory activities. This study investigates the healing effect of AE extract in comparison with SSD in second degree burn wounds. METHODS: Fort eight female Wistar rats (220±20 g) were divided into four groups. Standard second degree burn wounds were induced on the back of their necks. One group was treated with SSD; two groups were treated with AE cream at concentrations of 10% (AE10) and 20% (AE20) and the control group which received no treatment. The duration of treatment was 28 days. RESULTS: This study revealed that AE and SSD noticeably improved re-epithelization, fibroblasts proliferation, and collagen bundle synthesis and had a noticeable anti-inflammatory effect compared with the control group. CONCLUSION: Results of the present study revealed that Arnebia euchroma herbal extract was an effective treatment for second degree burn wounds when compared with SSD.

Stereological study of the effects of vitamin E on testis structure in rats treated with para-nonylphenol.

This study was organized to see whether vitamin E, as a strong antioxidant, could affect the abnormalities of testis structure caused by para-nonylphenol (p-NP) during its development. A total of 32 female Wistar rats after mating were divided into four groups (n = 8): control, vitamin E (100 mg kg(-1) per day), p-NP (250 mg kg(-1) per day) and p-NP + vitamin E. The rats were treated from the seventh day of pregnancy till the twenty-first day. After weaning, the male pups were divided into the same groups and were treated orally for 90 days. Finally, the right testis was fixed, processed, stained and studied using stereological methods. The weight and volume of testis, volume of seminiferous tubules and its diameter, thickness of the basement membrane, height of the germinal epithelium, total number of types A and B spermatogonia, spermatocyte, spermatid and Sertoli cells were significantly reduced in p-NP group when compared with other groups. Co-administration of vitamin E and p-NP compensated for the adverse effects of p-NP on the above parameters. In addition, treatment with only vitamin E caused a significant increase in diameter, basement membrane thickness and height of germinal epithelium, number of spermatogonia and spermatocytes. Co-administration of vitamin E with p-NP could prevent the adverse effects of p-NP on the testis structure during its development.