Effects of Polylactide Copolymer Implants and Platelet-Rich Plasma on Bone Regeneration within a Large Calvarial Defect in Sheep

The aim of this study was to verify whether L-lactide/DL-lactide copolymer 80/20 (PLDLLA) and platelet-rich plasma (PRP) trigger bone formation within critical-sized calvarial defects in adult sheep ( = 6). Two craniectomies, each ca. 3 cm in diameter, were created in each animal. The first craniectomy was protected with an inner polylactide membrane, filled with PRP-polylactide granules, and covered with outer polylactide membrane. The second control craniectomy was left untreated. The animals were euthanized at 6, 7, 17, 19, 33, and 34 weeks after surgery, and the quality and the rate of reossification were assessed histomorphometrically and microtomographically.The study demonstrated that application of implantsmade of PLDLLA 80/20 combined with an osteopromotive substance (e.g., PRP) may promote bone healing in large calvarial defect in sheep. These promising proof-of-concept studies need to be verified in the future on a larger cohort of animals and over a longer period of time in order to draw definitive conclusions

Rat spleen in the course of Babesia microti invasion: histological and submicroscopic studies

The course of babesiosis in humans is characterized by various intensity levels − from a subclinical level to the severe one − associated with multiple organ failure, which leads to death. The aim of this study was to evaluate the effect of 21-day and 6-month invasion of B. microti on Wistar rats spleen. Histological changes in the rats’ spleen were characterized by swelling of splenic tissue, especially the tissue adjacent to the capsule. In the structure of the white pulp in some rats, high concentrations of lymphocytes occurred. The boundary between the white pulp and red pulp was blurred. In the red pulp structure of rats, a lot of macrophages and extracellular deposits of bilirubin were present. The submicroscopic studies showed that the nuclear matrix was slightly shrunken. In the red pulp fragments of the damaged cells were located in the intercellular spaces. Near these areas, many thrombocytes were visible. The ultrastructural observation also revealed thickened endoplasmic reticulum membranes, local cellular swelling filled with amorphous substance, and digested erythrocytes. B. microti invasion affects the splenic morphology and ultrastructure in rats. The immunological hyperactivity and signs of inflammation indicate an important role of spleen in a fight against parasites

Administration of Cyclosporine A in Pregnant Rats - the Effect on Blood Pressure and on the Glomerular Number in Their Offspring

Background/Aims: Cyclosporine A (CsA) is a commonly used immunosuppressive agent. In some patients treatment with CsA has to be continued during pregnancy. The aim of the study was to assess in an experimental model whether the exposure to CsA during fetal life influences the number and volume of glomeruli, kidney function and blood pressure in the offspring. Methods: Eight pregnant female Sprague-Dawley rats were allocated to 2 treatment regimens: with CsA or solvent. Blood pressure was measured in the offspring at 7 and 11 weeks of age and albuminuria was determined at 11 weeks of age. In the kidney the number and mean volume of glomeruli was assessed using stereological methods. Results: In the offspring of pregnant rats treated with CsA the number of glomeruli was significantly lower and the mean volume of glomeruli was higher when compared to the offspring of pregnant rats receiving solvent. Systolic and diastolic blood pressures as well as albuminuria were significantly higher in the offspring of mothers treated with CsA during gestation compared to the offspring from the control group. Conclusions: Exposure of rats to CsA during fetal life impairs kidney development, thus potentially predisposing to chronic kidney disease and hypertension in the adult life

Modulatory effect of long-term treatment with escitalopram and clonazepam on the expression of anxiety-related neuropeptides:neuromedin U, neuropeptide S and their receptors in the rat brain

BACKGROUND: Newly identified multifunctional peptidergic modulators of stress responses: neuromedin U (NMU) and neuropeptide S (NPS) are involved in the wide spectrum of brain functions. However, there are no reports dealing with potential molecular relationships between the action of diverse anxiolytic or antidepressant drugs and NMU and NPS signaling in the brain. The present work was therefore focused on local expression of the aforementioned stress-related neuropeptides in the rat brain after long-term treatment with escitalopram and clonazepam. METHODS: Studies were carried out on adult, male Sprague-Dawley rats that were divided into 3 groups: animals injected with saline (control) and experimental individuals treated with escitalopram (at single dose 5 mg/kg daily), and clonazepam (at single dose 0.5 mg/kg). All individuals were sacrificed under anaesthesia and the whole brain excised. Total mRNA was isolated from homogenized samples of amygdala, hippocampus, hypothalamus, thalamus, cerebellum and brainstem. Real time-PCR method was used for estimation of related NPS, NPS receptor (NPSR), NMU, NMU and receptor 2 (NMUR2) mRNA expression. The whole brains were also sliced for general immunohistochemical assessment of the neuropeptides expression. RESULTS: Chronic administration of clonazepam resulted in an increase of NMU mRNA expression and formation of NMU-expressing fibers in the amygdala, while escitalopram produced a significant decrease in NPSR mRNA level in hypothalamus. Long-term escitalopram administration affects the local expression of examined neuropeptides mRNA in a varied manner depending on the brain structure. CONCLUSIONS: Pharmacological effects of escitalopram may be connected with local at least partially NPSR-related alterations in the NPS/NMU/NMUR2 gene expression at the level selected rat brain regions. A novel alternative mode of SSRI action can be therefore cautiously proposed

Escitalopram alters local expression of noncanonical stress-related neuropeptides in the rat brain via NPS receptor signaling

Background Neuropeptide S (NPS) is a multifunctional regulatory factor that exhibits a potent anxiolytic activity in animal models. However, there are no reports dealing with the potential molecular relationships between the anxiolytic activity of selective serotonin reuptake inhibitors (SSRIs) and NPS signaling, especially in the context of novel stress-related neuropeptides action. The present work therefore focused on gene expression of novel stress neuropeptides in the rat brain after acute treatment with escitalopram and in combination with neuropeptide S receptor (NPSR) blockade. Methods Studies were carried out on adult, male Sprague–Dawley rats that were divided into five groups: animals injected with saline (control) and experimental rats treated with escitalopram (at single dose 10 mg/kg daily), escitalopram and SHA-68, a selective NPSR antagonist (at a single dose of 40 mg/kg), SHA-68 alone and corresponding vehicle (solvent SHA-68) control. To measure anxiety-like behavior and locomotor activity the open field test was performed. All individuals were killed under anaesthesia and the whole brain was excised. Total mRNA was isolated from homogenized samples of the amygdala, hippocampus, hypothalamus, thalamus, cerebellum, and brainstem. Real-time PCR was used for estimation of related NPS, NPSR, neuromedin U (NMU), NMU receptor 2 (NMUR2) and nesfatin-1 precursor nucleobindin-2 (NUCB2) gene expression. Results Acute escitalopram administration affects the local expression of the examined neuropeptides mRNA in a varied manner depending on brain location. An increase in NPSR and NUCB2 mRNA expression in the hypothalamus and brainstem was abolished by SHA-68 coadministration, while NMU mRNA expression was upregulated after NPSR blockade in the hippocampus and cerebellum. Conclusions The pharmacological effects of escitalopram may be connected with local NPSR-related alterations in NPS/NMU/NMUR2 and nesfatin-1 gene expression at the level of selected rat brain regions. A novel alternative mode of SSRI action can be therefore cautiously proposed

A Novel Improved Thromboembolism-Based Rat Stroke Model That Meets the Latest Standards in Preclinical Studies

The animal thromboembolic model of ischemia perfectly mimics human ischemic stroke which remains the leading cause of disability and mortality in humans. The development of new treatment strategies was therefore imperative. The purpose of this study is to improve the thromboembolic stroke model in rats in order to design experiments that use motor tests, and are in accordance with the 3R principles to prevent complications and maintain the same size of the infarct repeatedly. Tail vein dye application, a protective skull mask and a stress minimization protocol were used as additional modifications to the animal stroke model. These modifications significantly minimized the pain and stress severity of the procedures in this model. In our experimental group of Long-Evans rats, a photo-induced stroke was caused by the application of a photosensitive dye (Rose Bengal) activated with white-light irradiation, thus eliminating the need to perform a craniotomy. The animals’ neurological status was evaluated using a runway elevated test. Histological examination of the brain tissue was performed at 12, 24 and 48 h, and seven days post-stroke. Tissue examination revealed necrotic foci in the cortex and the subcortical regions of the ipsilateral hemisphere in all experimental groups. Changes in the area, width and depth of the necrotic focus were observed over time. All the experimental groups showed motor disturbances after stroke survival. In the proposed model, photochemically-induced stroke caused long-term motor deficits, showed high reproducibility and low mortality rates. Consequently, the animals could participate in motor tests which are particularly suitable for assessing the efficacy of neuro-regenerative therapies, while remaining in line with the latest trends in animal experimental design

After Ischemic Stroke, Minocycline Promotes a Protective Response in Neurons via the RNA-Binding Protein HuR, with a Positive Impact on Motor Performance

Ischemic stroke is the most common cause of adult disability and one of the leading causes of death worldwide, with a serious socio-economic impact. In the present work, we used a new thromboembolic model, recently developed in our lab, to induce focal cerebral ischemic (FCI) stroke in rats without reperfusion. We analyzed selected proteins implicated in the inflammatory response (such as the RNA-binding protein HuR, TNFa, and HSP70) via immunohistochemistry and western blotting techniques. The main goal of the study was to evaluate the beneficial effects of a single administration of minocycline at a low dose (1 mg/kg intravenously administered 10 min after FCI) on the neurons localized in the penumbra area after an ischemic stroke. Furthermore, given the importance of understanding the crosstalk between molecular parameters and motor functions following FCI, motor tests were also performed, such as the Horizontal Runway Elevated test, CatWalk (TM) XT, and Grip Strength test. Our results indicate that a single administration of a low dose of minocycline increased the viability of neurons and reduced the neurodegeneration caused by ischemia, resulting in a significant reduction in the infarct volume. At the molecular level, minocycline resulted in a reduction in TNFa content coupled with an increase in the levels of both HSP70 and HuR proteins in the penumbra area. Considering that both HSP70 and TNF-a transcripts are targeted by HuR, the obtained results suggest that, following FCI, this RNA-binding protein promotes a protective response by shifting its binding towards HSP70 instead of TNF-a. Most importantly, motor tests showed that reduced inflammation in the brain damaged area after minocycline treatment directly translated into a better motor performance, which is a fundamental outcome when searching for new therapeutic options for clinical practice