Development of a rat model of knee osteoarthritis by a combination of monoiodoacetate and streptozotocin / Desarrollo de un modelo de rata de osteoartritis de rodilla por una combinación de monoiodoacetato y estreptozotocina

SUMMARY: Osteoarthritis (OA) caused by ageing joints or as a secondary complication of diabetes is a common health problem. We sought to develop an animal model of OA induced by a combination of the chondrocyte glycolytic inhibitor mono-iodoacetate (MIA) and streptozotocin (STZ), the agent that induces diabetes mellitus. We then hypothesized that the extent of damages to the knee joint induced by this model can be greater than OA induced by either MIA or STZ. Rats were either injected with MIA (model 1) or STZ (model 2) or both agents (model 3). After 8 weeks, harvested tissues from the knee joint of these groups were examined using scanning and transmission electron microscopy. In addition, blood samples were assayed for tumor necrosis factor alpha (TNF-α) and interleukin -6 (IL-6) that are known to be modulated in OA and diabetes. Compared to control group, substantial damages to the articular cartilage of the knee joint were observed in the three models with the severest in model 3. In addition, rats in model 3 showed significant (P<0.0001) increase in TNF-α and IL-6 compared to model 1 and 2. Thus, we have developed a new model of knee OA in rats that mimics a type of OA that is common among elderly people who have both, "ageing" joints and diabetes.

RESUMEN: La osteoartritis (OA) es un problema generalizado de salud a causa de un envejecimiento de las articulaciones, o bien de una complicación secundaria de la diabetes. El objetivo de este estudio fue desarrollar un modelo animal de OA inducido por una combinación dos drogas, un inhibidor de los condrocitos glucolíticos, el mono-iodoacetato (MIA), y la estreptozotocina (STZ), agente que induce la diabetes mellitus. Se consideró como hipótesis que el alcance de los daños a la articulación de la rodilla inducida por este modelo puede ser mayor que la OA inducida por MIA o STZ. Las ratas fueron inyectadas con MIA (grupo 1) o STZ (grupo 2) o ambos agentes (grupo 3). Se extrajeron muestras de la articulación de la rodilla de estos grupos al término de 8 semanas, y se examinaron mediante microscopía electrónica de barrido y de transmisión. Además, se analizaron muestras de sangre para el factor de necrosis tumoral alfa (TNF-α) e interleucina-6 (IL-6), que están moduladas en OA y en la diabetes. En comparación con el grupo control, se observaron daños sustanciales en el cartílago articular de la articulación de la rodilla en los tres modelos, encontrándose los daños más severos en el grupo 3. Además, las ratas del grupo 3 mostraron un aumento significativo (P <0,0001) de los niveles de TNF-α e IL- 6, en comparación con los grupos 1 y 2. Hemos desarrollado un nuevo modelo de OA de rodilla en ratas que imita un tipo de OA el cual, además de la diabetes, es común entre las personas mayores con un nivel importante de daño en las articulaciones.

Oxidative damage in embryo and placenta of streptozotocin-induced diabetic rats

Maternal type 1 diabetes is associated with an increased risk for fetal malformations. The mechanism by which diabetes caused teratogenic disorders is not fully known. Previous studies have demonstrated that many teratogenic diabetic cases were related to free radical oxygen species. This study was conducted to evaluate the effect of maternal diabetes on both embryo and placenta by estimating the oxidative and DNA damage in embryo and placenta of diabetic mellitus- induced rats. The possible role of olive leaves extract of Olea europaea [O. europaea] plant in repairing the damage was also assessed. Diabetes mellitus was induced by streptozotocin [STZ] by a single intraperitoneal injection [35 mg/kg b wt]. O.europaea leaves water extract was administered orally [550 mg/ l00g b wt/ day] for 5days before pregnancy and 18 days after. Malondealdehyde [MDA] level, glutathione peroxidase [Gpx] and superoxide dismutase [SOD] activities and glycogen concentration were measured in term embryo and placenta homogenates of diabetic and control rats. Moreover, the evaluation of DNA damage was carried out by the Alkaline Comet Assay using embryos and placentas taken from STZ-induced diabetic and control pregnant rats. the results showed an elevation in MDA level of the diabetic groups of both embryo and placenta compared to that of the control. This was accompanied by reduction in Gpx and SOD activities indicating oxidative damage. Glycogen level was reduced in diabetic groups of embryo and placenta. Both oxidative and hyperglycemic status were improved in the groups treated with olive leaves water extract. The percentage of tail DNA and tail moment values were also higher in both embryo and placenta of the diabetic -induced rats. DNA damage seems to be partly ameliorated in groups treated with O. europaea leaves water extract. This study indicated that maternal hyperglycemic condition in diabetic- induced pregnant rats could generate oxidative and DNA damage to embryo and placenta that could be ameliorated by oral doses of olive leaves water extract

Agmatine Improves Cognitive Dysfunction and Prevents Cell Death in a Streptozotocin-Induced Alzheimer Rat Model

PURPOSE: Alzheimer's disease (AD) results in memory impairment and neuronal cell death in the brain. Previous studies demonstrated that intracerebroventricular administration of streptozotocin (STZ) induces pathological and behavioral alterations similar to those observed in AD. Agmatine (Agm) has been shown to exert neuroprotective effects in central nervous system disorders. In this study, we investigated whether Agm treatment could attenuate apoptosis and improve cognitive decline in a STZ-induced Alzheimer rat model. MATERIALS AND METHODS: We studied the effect of Agm on AD pathology using a STZ-induced Alzheimer rat model. For each experiment, rats were given anesthesia (chloral hydrate 300 mg/kg, ip), followed by a single injection of STZ (1.5 mg/kg) bilaterally into each lateral ventricle (5 microL/ventricle). Rats were injected with Agm (100 mg/kg) daily up to two weeks from the surgery day. RESULTS: Agm suppressed the accumulation of amyloid beta and enhanced insulin signal transduction in STZ-induced Alzheimer rats [experimetal control (EC) group]. Upon evaluation of cognitive function by Morris water maze testing, significant improvement of learning and memory dysfunction in the STZ-Agm group was observed compared with the EC group. Western blot results revealed significant attenuation of the protein expressions of cleaved caspase-3 and Bax, as well as increases in the protein expressions of Bcl2, PI3K, Nrf2, and gamma-glutamyl cysteine synthetase, in the STZ-Agm group. CONCLUSION: Our results showed that Agm is involved in the activation of antioxidant signaling pathways and activation of insulin signal transduction. Accordingly, Agm may be a promising therapeutic agent for improving cognitive decline and attenuating apoptosis in AD.

Effect of Bilirubin on Triglyceride Synthesis in Streptozotocin-Induced Diabetic Nephropathy

We aimed to elucidate the effect of bilirubin on dyslipidemia and nephropathy in a diabetes mellitus (DM) type I animal model. Sprague-Dawley rats were separated into control, DM, and bilirubin-treated DM (Bil) groups. The Bil group was injected intraperitoneally with 60 mg/kg bilirubin 3 times per week and hepatoma cells were cultured with bilirubin at a concentration of 0.3 mg/dL. The Bil group showed lower serum creatinine levels 5 weeks after diabetes onset. Bilirubin treatment also decreased the amount of mesangial matrix, lowered the expression of renal collagen IV and transforming growth factor (TGF)-beta1, and reduced the level of apoptosis in the kidney, compared to the DM group. These changes were accompanied by decreased tissue levels of hydrogen superoxide and NADPH oxidase subunit proteins. Bilirubin decreased serum total cholesterol, high-density lipoprotein cholesterol (HDL-C), free fatty acids, and triglycerides (TGs), as well as the TG content in the liver tissues. Bilirubin suppressed protein expression of LXRalpha, SREBP-1, SCD-1, and FAS, factors involved in TG synthesis that were elevated in the livers of DM rats and hepatoma cells under high-glucose conditions. In conclusion, bilirubin attenuates renal dysfunction and dyslipidemia in diabetes by suppressing LXRalpha and SREBP-1 expression and oxidative stress.

Neurobiological effect of 7-nitroindazole, a neuronal nitric oxide synthase inhibitor, in experimental paradigm of Alzheimer’s disease.

Nitric oxide plays a role in a series of neurobiological functions, underlying behaviour and memory. The functional role of nNOS derived nitric oxide in cognitive functions is elusive. The present study was designed to investigate the effect of specific neuronal nitric oxide synthase inhibitor, 7-nitroindazole, against intracerebroventricular streptozotocin-induced cognitive impairment in rats. Learning and memory behaviour was assessed using Morris water maze and elevated plus maze. 7-nitroindazole (25 mg/kg, ip) was administered as prophylactically (30 min before intracerebroventricular streptozotocin injection on day 1) and therapeutically (30 min before the assessment of memory by Morris water maze on day 15). Intracerebroventricular streptozotocin produced significant cognitive deficits coupled with alterations in biochemical indices.These behavioural and biochemical changes were significantly prevented by prophylactic treatment of 7-nitroindazole. However, therapeutic intervention of 7-nitroindazole did not show any significant reversal. The results suggests that 7-nitroindazole can be effective in the protection of dementiainduced by intracerebroventricular streptozotocin only when given prophylactically but not therapeutically.

Diabetes-induced prefrontal nissl substance deficit and the effects of neem-bitter leaf extract treatment / Diabetes inducida por déficit prefrontal de sustancia de nissl y el efecto del tratamiento con extracto de hoja de neem amargo

Cognitive dysfunction is reportedly associated with poorly-managed diabetes mellitus. In this study, we report the effect of oral treatment with combined leaf extract (CLE) of neem and bitter leaf on the prefrontal cortex of diabetic Wistar rats. Adult male Wistar rats were randomized to one of the following groups: control, diabetic (STZ-induced), STZ + CLE, STZ + metformin and CLE only. At euthanasia, paraffin sections of the prefrontal cortex were stained with cresyl fast violet; while malondialdehyde (MDA) and glutathione peroxidase (GPx) were assayed in prefrontal homogenates. Oral CLE produced normoglycemia in the treated hyperglycaemic rats. Besides, Nissl-stained prefrontal sections showed no morphologic deficits in all the groups except the untreated diabetic rats. In the latter, there was weak Nissl staining, while prefrontal MDA was significantly high at euthanasia, compared with the control and CLE-treated rats (P<0.05). This study showed that untreated diabetes mellitus is associated with prefrontal Nissl body deficit and oxidative stress in Wistar rats. The absence of these deficits in CLE-treated rats suggests a neuroprotective effect of the extract in streptozotocin-induced diabetic rats. This may improve the cognitive function of the prefrontal cortex in diabetes mellitus.

La disfunción cognitiva es presuntamente asociada con un mal manejo de la diabetes mellitus. En este estudio, se presenta el efecto del tratamiento oral combinado con extracto de hoja (CLE) de hoja de neem amarga sobre la corteza prefrontal de ratas Wistar con diabetes. Las ratas Wistar adultas fueron asignadas al azar a uno de los siguientes grupos: control, diabetes (STZ inducida), STZ + CLE, STZ + metformina y CLE. Después de la eutanasia, los cortes de parafina de la corteza prefrontal se tiñeron con violeta de cresil rápido, mientras que el malondialdehído (MDA) y la glutatión peroxidasa (GPx) fueron analizadas en homogenizados prefrontales. El CLE produce normoglucemia en las ratas hiperglucémicas tratadas. Además, las secciones prefrontales teñidas para Nissl no muestran ningún déficit morfológico en todos los grupos excepto en las ratas diabéticas sin tratamiento. En este último caso, hubo una tinción de Nissl débil, mientras que la MDA prefrontal fue significativamente más alta en comparación con los grupos de ratas control y las tratadas con CLE (p <0,05). Este estudio mostró que la diabetes mellitus no tratada se asocia con déficit prefrontal de cuerpos de Nissl y estrés oxidativo en ratas Wistar. La ausencia de estos déficits en las ratas tratadas CLE, sugiere un efecto neuroprotector del extracto en ratas diabéticas inducidas por estreptozotocina. Esto puede mejorar la función cognitiva de la corteza prefrontal en la diabetes mellitus.

Effects of Danae racemosa on testosterone hormone in experimental diabetic rats

Danae racemosa is a strong antioxidant and antioxidants have significant effects on spermatogenesis, sperm biology and oxidative stress, and changes in antioxidant capacity are considered to be involved in the pathogenesis of chronic diabetes mellitus. Because STZ causes testicular dysfunction and degeneration under situations of experimentally induced diabetes in animal models, we aimed in this study Danae racemosa's effects in decreasing the harmful effects of STZ on testicular and sperm functions b. Male Wistar rats [n = 40] were allocated into four groups: Group 1a: Control rats given 0.5 ml of 20% glycerol in 0.9% normal saline. Group 1b: Control rats given 0.5ml of 0.5 ml citrate buffer [pH4.0].Group 2: streptozotocin [STZ] treated rats. Group 3: rats given danae racemosa 400 mg/kg [gavage]. Group 4: STZ treated rats given danae racemosa 400 mg/kg [gavage]. Animals were kept in standard conditions. At 28 days after inducing diabetics, 5 mL blood was collected for measuring testosterone. Total serum testosterone increased significantly in the group treated with danae racemosa [p < 0.05] compared with control groups. Testis weights in the diabetic groups decreased significantly in comparison with controls [p < 0.05]. Danae racemosa had a significant protective effect on the diabetes-induced deteriorations in serum total testosterone, by reducing the levels of reactive oxygen species in serum. Therefore, it could be effective for maintaining healthy in diabetic rats

[Comparison of the toxic effects related to experimental induction of diabetes by Alloxam and Strepytozotocin in rats]

Both Alloxan and Streptozotocin [STZ] are selective substances used to induce experimental diabetes, because they have an important property to damage beta-cells that secrete insulin in Pancreas. But their use has some problems; they have toxic effects on important body organs and also cause some necrosis in tissue. The aim of this study was to clarify the best way of administration, so we examine the acute way which includes giving a single high dose of both the previous materials. Indeed, we also examine a gradual way that includes giving the substances in repeated low doses with intervals last for several days. We measured the changes of weight, the elevation in blood sugar and the mortality for both previous ways. Indeed, a histological study was performed for important body's organ following the administration of the mentioned materials in the gradual way. The findings indicated clearly that the gradual way for both Alloxan and STZ was more safe and effective than acute administration. STZ was best from Alloxan especially in the gradual way concerning the inducing of diabetes and the low mortality. According to these results we recommend to use STZ with the gradual way in the researches that require experimental diabetic animals

Pioglitazone, a ppar-gamma ligand inhibited the nicotinamide streptozotocin induced sperm abnormalities in type-2 diabetic wistar rats

Oxidative stress due to chronic hyperglycemia is known to cause infertility in diabetic patients. Pioglitazone [PIO], a PPAR-gamma ligand is known to possess the antioxidant property however, its role on the oxidative stress mediated germinal damage in Type-2 diabetes mellitus [T2DM] is poorly studies in the literature. In this study, the effect of PIO [1 and 10 mg/kg, p.o. daily for 4 weeks] was evaluated against the nicotinamide [NA-230 mg/kg, ip] and streptozotocin [STZ-65 mg/kg, ip] induced changes in sperm abnormality and antioxidant status in Wistar rats. The effect of PIO on the male reproductive cells was studied by determining the sperm shape abnormality and sperm count in the experimental T2DM. The antioxidant profile was studied by estimating the serum lipid peroxidation [LPO], glutathione [GSH] and glutathione peroxidase [GPx] levels. The higher dose of PIO [10 mg/kg] significantly [P<0.001] reduced the sperm shape abnormality and increased the sperm count [P<0.00l], besides decreasing [P<0.05] the LPO and enhancing the GSH and GPx levels in the diabetic rats. However, the lower dose of PIO [1 mg/kg] produced inhibitory [P<0.05] effect against the changes induced by T2DM in the sperm shape abnormality and GSH levels. alpha tocopherol tested as alpha standard antioxidant agent prevented [P<0.001] the NA-STZ mediated alterations in sperm abnormalities and antioxidant enzyme levels. The results suggest that the inhibitory effect of PIO against the 12DM induced sperm abnormality could be related to its antioxidant property

Altered kidney morphology and enzymes in streptozotocin induced diabetic rats / Morfología y enzimas renales alteradas en ratas con diabetes inducida por estreptozotocin

We studied the effects of streptozotocin (STZ)-induced diabetes on kidney morphology, anatomy, architecture and on the activities of aminotransferases (AST and ALT), alkaline phosphatase (ALP) and pseudocholinesterase (PChE) in albino rats. The aim of the study was to investigate the association between diabetic kidney complications and kidney enzyme alterations. This study was performed in the Department of Anatomy and Institute of Pharmaceutical Sciences, Baqai Medical University, Karachi and Pathology department of College of Physicians & Surgeons (CPSP) Pakistan in 2007-08. Diabetes was induced by a single dose of STZ (45 mg/kg, b.w.) given intraperitoneally in sodium citrate buffer at pH 4.5. Eighty (80) albino rats were divided into five groups: control (A) and STZ treated (B, C, D, and E) which were sacrificed 2, 4, 6 and 8 weeks post treatment respectively. Histopathology of kidney showed lesions similar to human glomerulosclerosis, glomerular membrane thickening, arteriolar hyalinization and tubular necrosis. Increased levels of aspartate aminotransferase (AST), alanine aminotransferase (ALT), alkaline phosphatase (ALP) and pseudocholinesterase (PChE) were observed in the kidney. It seems that the diabetic complications in the kidney are likely to be associated with alterations in enzyme levels.

Se estudiaron los efectos de la diabetes inducida por estreptozotocin (STZ) sobre la morfología, anatomía, arquitectura y sobre las actividades de aminotransferasas (ALT y AST), fosfatasa alcalina (ALP) y pseudocolinesterasa (PChE) en los riñones de ratas albinas. El objetivo del estudio fue investigar la asociación entre las complicaciones renales diabéticas y la alteración de las enzimas renales. Este estudio se realizó en el Departamento de Anatomía y el Instituto de Ciencias Farmacéuticas, Universidad de Medicina Baqai, Karachi y el departamento de Patología de Colegio de Médicos y Cirujanos (CPSP) Pakistán entre el 2007-2008. La diabetes fue inducida por una dosis única de STZ (45 mg / kg de peso corporal) administrada por vía intraperitoneal en tampón de citrato de sodio a pH 4.5. Ochenta (80) ratas albinas fueron divididas en cinco grupos: control (A) y STZ tratados (B, C, D y E), que se sacrificaron a las 2, 4, 6 y 8 semanas después del tratamiento, respectivamente. La histopatología del riñón mostró lesiones similares a la glomeruloesclerosis en humanos, engrosamiento de la membrana glomerular, hialinización arteriolar y necrosis tubular. Aumento de los niveles de aspartato aminotransferasa (AST), alanina aminotransferasa (ALT), fosfatasa alcalina (ALP) y pseudocolinesterasa (PChE) fueron observados en el riñón. Parece que las complicaciones de la diabetes en el riñón están directamente asociadas con alteraciones en los niveles de las enzimas.

Preliminary studies on antihyperglycemic effect of aqueous extract of Brassica nigra (L.) Koch in streptozotocin induced diabetic rats.

In the streptozotocin induced diabetic rats treated separately with aqueous, ethanol, acetone and chloroform extracts of the seeds of B. nigra, the increase in serum glucose value between 0 and 1 hr of glucose tolerance test (GTT) was the least (29 mg/dl) in aqueous extract treated animals while it was 54, 44 and 44 mg/dl with chloroform, acetone and ethanol extracts respectively. In further studies carried out with aqueous extract, the effective dose was found to be 200 mg/kg body weight in GTT. Administration of 200 mg/kg body weight of aqueous extract to diabetic animals daily once for one month brought down fasting serum glucose (FSG) levels while in the untreated group FSG remained at a higher value. In the treated animals the increase in glycosylated hemoglobin (HbA1c) and serum lipids was much less when compared with the levels in untreated diabetic controls. These findings suggest that further studies with the aqueous extract of B. nigra seeds on its antidiabetic activity would be useful.

Effect of Aegle marmelos Correa. (Bael) fruit extract on tissue antioxidants in streptozotocin diabetic rats.

A study was undertaken to evaluate the anti-lipid peroxidative activity of an aqueous extract of A. marmelos fruits (AMFEt) in streptozotocin diabetic rats in heart and pancreas. Oral administration of AMFEt for 30 days (125 and 250 mg kg(-1) body weight twice daily) produced a significant decrease in the elevated levels of peroxidation products, viz. thiobarbituric acid reactive substances and hydroperoxides in the tissues of diabetic rats. The depressed activities of superoxide dismutase, catalase and glutathione peroxidase and lowered glutathione content in the heart and pancreas of diabetic rats were found to increase on treatment with AMFEt. AMFEt at a dose of 250 mg kg(-1) was more effective than glibenclamide (300 microg kg(-1)) and both reversed all the values significantly. Thus AMFEt exhibits anti-oxidative activity in streptozotocin diabetic rats.

optimum dose of nicotinamide for protection of pancreatic Beta-cells against the cytotoxic effect of streptozotocin in albino rat

The natural course of Insulin Dependent Diabetes Mellitus is characterized by progressive destruction of insulin producing Beta-cells of the pancreas resulting from an autoimmune process. The toxic effect of some Beta-cells toxins like streptozotocin [used to produce animal models of IDDM] has been associated with the oxidative stress due to enhanced DNA repair and NAD depletion in damaged Beta-cells. This activity of streptozotocin has been prevented with the use of nicotinamide. A light microscopic study was designed to determine the optimum dose of nicotinamide required for protection of pancreatic Beta cells against the toxicity of streptozotocin. 35 adult male albino rats were divided into five equal groups A, B, C, D and E. the duration of study was 14 days. The animals in experimental groups C, D and E received a single intraperitoneal injection of nicotinamide 250 mg/Kg, 350 mg/Kg and 500 mg/Kg respectively on day one. Animals in group A and B acted as normal control and diabetic control respectively. All the animals except those in group A received simultaneous injection of streptozotocin 32 mg /Kg body weight intraperitoneally in a single dose. Fasting blood glucose was assessed and the animals weighed before starting the treatment, after 48 hours and at the end of the experimental period. Histological studies were carried out at the end of the study period. The blood glucose level and the final body weight of the animals in group C matched the values in diabetic control. Histologically the pancreas had generally reduced Beta-cells mass [P < 0.001] with altered morphology. The animals in group D showed impaired glucose tolerance at 48 hours but were normoglycaemic at the end of the study period. There was some loss of Beta-cells but a significant number of these cells [P < 0.05] showing normal morphology were saved. The animals in group E had normal number of P-cells having normal morphological features. The final body weight and fasting blood glucose of these animals matched the values in normal control [group A]. Conclusions: These data suggest that the optimum dose of nicotinamide in regard to prevention against the Beta-cytotoxic effect of streptozotocin in albino rat is 500 mg/Kg body weight

Hepatic arylsulfatases A and B activities in streptozocin - induced diabetic rats

Reduction of sulfated glycosaminoglycans [GAG] in the liver and kidney of streptozocin-induced diabetic rats has been attributed to lowered synthesis and perhaps higher degradation of such compounds in these organs. To measure hepatic lysosomal arylsulfatases A and B, [the enzymes responsible for the removal of sulfate groups from GAG], in starved and streptozocin-induced diabetic rats. Diabetes was induced by streptozocin injection [40 mg per kg body weight] through the caudal vein in rats. After two weeks, the livers were removed and homogenized. Activities of arylsulfatases A and B were measured and compared with those of the liver homogenates from healthy and starved rats. The activity of liver arylsulfatase A in starved diabetic rats increased 2.15 fold as compared with normal starved animals, while that of fed diabetic rats was 3.16 fold higher than their respective control group. Increases of 1.70 and 1.94 fold in specific activities of arylsulfatase B was noticeable in the livers of diabetic animals under fed and starved conditions, respectively. It appears that increased hydrolysis of sulfated GAG by liver lysosomal arylsulfatases A and B in streptozocin-induced diabetic rats may be among the contributing factors in the reduction of such sulfated compounds in this tissue

Morhological and quantitative analysis of the neurons of myenteric plexus of the cecum of streptozotocin-induced diabetic rats

The purpose of this work was to study the neurons of the myenteric plexus of the cecum of rats with chronic streptozotocin-induced diabetes. We used four experimental groups of animals. In groups D2 and D8 animals were killed two and eight months, respectively, after diabetes induction and groups C2 and C8 were used as controls. We carried out whole-mount preparations stained with Giemsa and NADH-diaphorase. We verified that the diabetes did not alter the shape and disposition of the myenteric ganglia; it provoked decrease on the neuronal density and increase on the incidence of weakly basophilic neurons. The effects of streptozotocin caused dilatation of the cecum still evidenced two months after induction, but no more observed on the eight months after induction. The smaller incidence of neuron in group D8 relative to group C8 was due to the early loss related to the drug toxicity and later to the aging in diabetic condition.