Animal Ethics Committee Research Articles

Prevention of mechanical valve thrombosis with a combination of the factor Xa inhibitor edoxaban plus aspirin in swine: intermediate results from the EdoValves project

Abstract Background Patients with an implanted mechanical heart valve (MHV) currently have a contraindication to the use of non-vitamin K antagonist oral anticoagulants (NOACs) based on the unfavorable results of the RE-ALIGN trial with dabigatran. We hypothesized that the combination of a Factor Xa inhibitor, acting upstream of thrombin generation, plus aspirin, may effectively prevent valve thrombosis, providing a much more friendly and viable alternative to vitamin K antagonists for such patients. We tested this in a swine model with a recently implanted mechanical valve prosthesis in the mitral position, combining the most unfavorable conditions promoting mechanical valve thrombosis, based on (a) the implanting position; (b) recent surgery; (c) the high thrombogenicity of the animal model. The study was approved by the Animal Ethics Committee of the University of Pisa and the Italian Ministry of Health and conducted according to Good Clinical Practices for Animal Experiments. Methods 17 farm male pigs, weighing ≈30 kg at the beginning, underwent cardiac surgery under general anesthesia and with extracorporeal circulation to receive a 25 mm or 27 mm bileaflet Corcym Bicarbon mechanical heart valve replacement in the mitral position. Pigs were given 1 mg/kg enoxaparin immediately after surgery and started with the oral daily administration of edoxaban 1 mg/kg (30 mg, then escalated to 60 mg with the animal growth) + aspirin 100 mg, both given as crushed tablets with a food pellet since the animal awakening from anesthesia. An initially planned warfarin control group was quickly abandoned because of the extreme difficulty of controlling warfarin activity. Animals were submitted to blood sampling and transthoracic echocardiography at least twice in the follow-up when uneventful for up to three months or in case of health deterioration. The autopsy was performed by a certified pathologist after each spontaneous death or procured sacrifice. Results (Figure 1): Excluding 5 peri-operative deaths, 12 animals survived surgery. Of those, 6 died within the second month: pulmonary edema occurred in 3 cases, due to prosthesis-mismatch in the growing animals; pulmonary infection occurred in 1 subject; heart failure due to valve thrombosis occurred in 2 cases. Six swine were full-term survivors in good health status up to the 3-month planned observational time. No hemorrhagic events were ever documented. Conclusion The combination of edoxaban + aspirin effectively avoided valve thrombosis events in 10 out of 12 (83%) analyzable animals and was safe from bleeding complications. The study is ongoing up to a predicted sample size of ≥20 animals. If further confirmed, these data may open the possibility of an alternative to vitamin K antagonists in humans with the fixed-dose drug combination here used, without the need for continuous anticoagulation monitoring and with expected greater safety.Figure 1

A comparative study of hypolipidemic effect of atorvastatin and Coriandrum sativum in triton-induced hyperlipidemic albino rat model

Background: Hyperlipidemia is the main factor responsible for coronary artery disease. Atorvastatin commonly used, as a competitive inhibitor of HMG-Co-A reductase, is known to reduce LDL and triglyceride levels and increase HDL levels. Coriandrum sativum (Coriander) belongs to the Apiaceae family and has hypolipidemic, antidiabetic, and antioxidant properties. Triton WR1339 has been used by several studies to induce hypercholesterolemia in animals which acts on both the synthesis and excretory phase. This study aimed to compare the hypolipidemic effect of Coriandrum sativum and atorvastatin in triton-induced hyperlipidemic rats. Methods: Following approval from the Institutional Animal Ethics committee, 36 Wistar albino rats were divided into 6 groups of 6 animals each. Triton 200mg IP was administered to all the groups except Group 1 (control). Groups 3 and 4 received Coriander extract 300mg/kg and Atorvastatin 80mg/kg orally with Triton. Groups 5 and 6 received Coriander extract and Atorvastatin respectively 22 hours later. Lipid profile was estimated 24 hours before, 24 and 48 hours after administering Triton. Results: In the synthesis phase, there is a statistically significant increase in the lipid levels in Groups 2, 5, and 6 compared to Groups 3 and, 4 indicating hypolipidemic effects of both Coriander extract and Atorvastatin. In the excretory phase, all groups have shown a decrease in lipid levels but a decrease in total cholesterol level in Group 5 compared to Group 3 was significant. Conclusions: Atorvastatin and Coriander extract both affect the lipid levels in the synthesis and excretory phase. Even though coriander cannot replace atorvastatin in the management of hyperlipidemia, it can be used as an adjuvant to atorvastatin to reduce the cost effects and adverse reactions caused by allopathic medicines.

8353 Impact of a plant-based protein diet on bone health and body composition in Sprague Dawley rats

Abstract Disclosure: M.M. Gomes: None. I.M. de Araújo: None. S.V. Rodriguez: None. F.R. Jorge: None. F.J. de Paula: None. Introduction:Vegetarian and vegan diets are associated with some health benefits; this has contributed to their growing popularity. Although studies remain limited, there is evidence that these patients may be at greater risk of reduced bone mineral density (BMD) and increased fracture risk. The general objective of this study was to analyze the impact of a high-protein, plant-based diet on bone tissue and body composition in male Sprague Dawley rats. Methodology: The study was approved by the Animal Ethics Committee. This study was done with 10 rats in the control group (CG), which received a standard AIN-93M diet; 11 in the normoprotein vegetarian group (NVG), with a normoprotein AIN-93 diet modified with pea protein and methionine; and 11 in the high-protein vegetarian group (HVG), with a modified AIN-93 diet with 40% pea protein and methionine. The dietary intervention lasted 8 weeks. The weight of the animals was measured weekly. Basal blood glucose levels were measured at baseline. At the end of the study, 6 rats from the CG, 6 rats from the NVG, and 5 rats from the HVG were subjected to a glucose tolerance test (GTT). The dual-energy X-ray absorptiometry (DXA) technique was applied in the baseline and final periods of the experiment to evaluate body composition, bone mineral content (BMC) and BMD. After sacrifice, bone strength was measured via a three-point flexion mechanical test on the tibia; brown adipose, epididymal, and retroperitoneal tissues, as well as the liver, were weighed on an analytical balance. Blood was used to measure circulatory levels of insulin, CTX, and P1NP (n=6/group). Results: Basal weight was similar between the groups, but the final weight was lower in the HVG as compared to the CG. In the DXA evaluation, the final lean mass was not different between the groups. On the other hand, total fat mass was lower in the HVG than in the CG and the NVG and the fat percentage was lower in the HVG than in the CG. The weight of epididymal adipose tissue was lower in the HVG than in the NVG, but retroperitoneal and brown adipose tissues were lower in HVG than CG. Liver weight was similar between groups. BMD was similar between groups; however, BMC was lower in the HVG than in the CG and the NVG. Bone strength was similar between groups. There was no difference in glycemia at baseline or during GTT at the eighth week. Serum levels of insulin, CTX, and P1NP were similar between groups. Discussion and Conclusion: A normoprotein vegetarian diet that includes all essential amino acids appears to have no negative impact on body composition or BMD in rats. On the other hand, a high-protein diet with plant protein appears to have a negative impact on bone mineral content, in addition to reducing the amount of body fat and weight. This finding expands on previous results by showing that high-protein diets have a negative impact on bone even when the protein is of plant origin. Presentation: 6/1/2024

Experimental Study of Hypolipidemic Effect of Manjal Noiku Kudineer

Hyperlipidemia, is a major stake for the development of serious diseases like atherosclerosis, Coronary Artery Diseases, Stroke. Siddha system of Medicine with the numerous combination of medicines has Manjal Noiku Kudineer(MNK), a Siddha poly herbal formulation indicated for Manjal Noi. This study deals with the Hypolipidemic activity of Manjal Noiku Kudineer on atherogenic diet induced hyperlipidemia in experimental Wistar Albino Rats Place of Study: The study was conducted at Arulmigu Kalasalingam College of Pharmacy, Virudhunagar. Methodology: The study protocol for MNK was reviewed and approved by Institutional Animal Ethics Committee (IAEC), Arulmigu Kalasalingam College of Pharmacy, Virudhunagar, with the IAEC number AKCP/IAEC/10/24-25. The experimental hyperlipidemic diet consists of well-pulverized 400 mg/kg cholesterol, 50 mg/kg cholic acid, and coconut oil. All groups other than the control group were fed the prepared atherogenic meal in lieu of the regular pellet diet. Group I served as normal control, Group II represented hyperlipidemic control rats given an atherogenic diet, Group III and IV represented test groups given MNK 200 mg/kg and MNK 400 mg/kg, respectively, and Group V represented standard-administered atorvastatin (10 mg/kg/day). All groups were compared, observations were noted and the observations were statistically evaluated. Results: The study shows the results of reduction in body weight, blood lipid profile, liver lipid profile of hyperlipidemic rats on treating with the study medicine, which possess hypolipidemic activity. Histopathologic images of rat liver also shown normal parenchyma and parenchyma with minimal necrosis and minimal inflammatory changes on treating with the study medicine MNK. Conclusion: The results stands as a support for MNK hypolipidemic activity, which in turn can be used as a antihyperlipidemic therapeutic agent from the Siddha System of Medicine.

P161 INVESTIGATION OF BIOACTIVE FLAVONOID SOURCED FROM CITRUS FRUITS ON ITS EFFECT UPON CARDIAC HYPERTROPHY: AN EXPERIMENTAL STUDY

Objectives: Cardiac hypertrophy is an adaptive process of the heart. It is a leading cause for heart failure, which in turn has a high mortality rate. Hesperidin (HES) is a bioactive flavanone glycoside with agonistic activity on Peroxisome Proliferator-Activated Receptor gamma (PPAR-γ), reported to be an inhibitor of cardiac hypertrophy signalling pathways. Therefore, we investigated the cardio-protective effect of this bioactive flavonoid in isoproterenol induced cardiac hypertrophy. Methods: The work was started after getting approval from the institutional animal ethics committee (IAEC). Male albino Wistar rats were divided into 6 groups (n=6) and administered isoproterenol (3 mg/kg/day s.c.) or hesperidin (200 mg/kg/day, p.o.) and isoproterenol or enalapril (30 mg/kg/day p.o.) and isoproterenol or a combination of hesperidin and enalapril for 28 days. On the 29 th day, rats were anaesthetized followed by cannulation of coronary artery to record hemodynamic parameters. Blood sample was collected and heart was excised to evaluate for parameters like biochemical, histopathological, ultrastructural and immunohistochemical studies. Results: Hesperidin significantly normalized heart weight/body weight ratio and cardiomyocyte area in the treated rats. Moreover, hesperidin improved cardiac function and attenuated pathological changes as seen in the heart tissue sections. A decline in the levels of cardiac injury markers (CK-MB, LDH) and inflammatory markers TNF-α, IL-6) was observed. Additionally, hesperidin suppressed oxidative stress and apoptosis along with an increased PPAR-γ expression. Conclusion: Hesperidin has a good potential for attenuation of cardiac hypertrophy and acts by virtue of its PPAR-γ agonistic, anti-inflammatory, anti-apoptotic and anti-oxidant properties.

Evaluation of Anxiolytic Activity of Angiotensin Receptor Blockers Using Actophotometer Test in Wistar Rats.

Introduction Anxiety disorders are common mental illnesses impacting quality of life, with current treatments like benzodiazepines andselective serotonin reuptake inhibitors (SSRIs)facing limitations due to side effects. Angiotensin receptor blockers (ARBs), used primarily for hypertension, have shown potential neuropsychiatric benefits, including anxiolytic effects. This study explores the anxiolytic effects of two ARBs, telmisartan and losartan, by evaluating locomotor activity in Wistar rats, aiming to identify new treatment options for anxiety through modulation of the renin-angiotensin system. Aim To evaluate the anxiolytic activity of telmisartan and losartan in experimental Wistar rats. Materials and methods The study was carried out afterconsent was obtained from the Institutional Animal Ethics Committee (IAEC). The rats were divided into four groups: group 1 (control group) received distilled water (2 ml/kg), group 2 (diazepam 2 mg/kg group) received diazepam (2 mg/kg), group 3 (telmisartan 5 mg/kg group) received telmisartan (5 mg/kg), and group 4 (losartan 5 mg/kg group) received losartan (5 mg/kg). The actophotometer test, which measures the locomotor activity levels of rats, was utilized to evaluate their anxiolytic activity. The percent decrease in locomotor activity was calculated for statistical evaluation. Results Our investigation found that the telmisartan 5 mg/kg and losartan 5 mg/kg groups had considerable anxiolytic activity (p<0.05) compared to the control group, and it was comparableto the diazepam 2 mg/kg (p>0.05) group. Conclusion The findings of our study indicate that ARBs, specifically telmisartan 5 mg/kg and losartan 5 mg/kg, exhibit potential anxiolytic effects, evidenced by a significant reduction in locomotor activity in the actophotometer test. These results imply that ARBs could be considered as possible therapeutic agents for anxiety, providing a new perspective on their use beyond traditional cardiovascular applications.

NOVEL ITACONATE DERIVATIVE ATTENUATES ISOPROTERENOL INDUCED CARDIOTOXICITY IN EXPERIMENTAL ANIMALS: A MOLECULAR PATHWAY INSIGHT

Objective: 4-octyl itaconate (4-OI) is a potential drug for the treatment of myocardial infarction on account of its potent anti-inflammatory, antioxidant and anti-apoptotic properties. Thus, we hypothesise that 4-OI may attenuate isoproterenol(ISO) induced cardiotoxicity by modulating several molecular pathways in Wistar albino rats. Design and method: After getting approval from animal ethics committee(278/IAEC/2021), the experimental work was started. ISO 5mg/kg s.c., was administered daily for 15 days. Adult male rats were randomly divided into 5 groups (n=6): normal control group, ISO control group, treatment control groups. 4-OI was administered i.p. at 3 different doses (2.5,5,10 mg/kg) to rats from day 1 and continued for 15 days along with ISO. On the 16th day, surgery was performed, right carotid artery was cannulated for recording of hemodynamic parameters. Blood was drawn using cardiac puncture and rats were then sacrificed, one portion of the heart was immediately stored in 10% formalin for histopathological evaluation and second portion was kept in -80 degree C for further studies. Serum was stored for biochemical, pro-inflammatory cytokines estimation (TNF, IL-6, IL1 beta) and cardiac injury markers (CKMB, LDH). Results: 4-OI improved cardiac function and attenuated pathological changes as seen microscopically in the H&amp;E stained heart tissue sections. On ultrastructural evaluation, 4-OI treatment (5, 10mg/kg) groups presented with lesser nuclear condensation and mitochondrial swelling, as compared to the ISO control group. A significant decline in the serum and tissue levels of cardiac injury markers (CK-MB, LDH) and inflammatory markers were observed in 4-OI (5,10mg/kg) treated rats, as compared to ISO control group. Additionally, 4-OI suppressed oxidative stress and apoptosis, along with an increased PPAR-gamma expression. Molecular pathway assessment revealed that at the dose of 10mg/kg, 4-OI reduced the expression of NFkappaB, IKK-beta and enhanced the expression of Nrf-2 and HO-1. On analysis of the inflammatory MAPK pathway, expression of the pro-survival kinase ERK was also increased while JNK and p38 protein expressions were reduced. Conclusions: 4-OI at a dose of 10mg/kg mitigates myocardial toxicity in ISO treated rats by maintaining morphological structure of heart tissue with oxidative stress profile and reduced inflammation

Effectiveness of Body Armor Against Shock Waves: Preventing Blast Injury in a Confined Space.

Introduction Blast injuries in modern society often occur owing to terrorist attacks in confined spaces, particularly in urban settings, indoors, and in vehicles, leading to significant damage. Therefore, it is important to focus on blast injuries in confined spaces rather than in conventional open-field experiments. Materials and methods We used an air-driven shock wave generator (blast tube) established indoors in 2017 and conducted basic research to potentially save the lives of patients with blast injuries. Under general anesthesia, pigs were divided into with body armor (BA) and without BA groups. The pigs were fixed in the measurement chamber with their dorsal chest directly exposed to the shock wave. The driving pressure was set at 3.0 MPa to achieve a mortality rate of approximately 50%. A generated shock wave was directly applied to the pigs. Comparisons were made between the groups with respect to cardiac arrest and survival, as well as apnea, bradycardia, and hypotension, which are the triad of blast lung. Autopsies were performed to confirm the extent of the organ damage. Statistical analysis was performed using Fisher's exact test, and statistical significance was set at p<0.05. The animal experimentation was conducted according to the protocol reviewed and approved by the Animal Ethics Committee of the National Defense Medical College Hospital (approval number 19041). Results Eight pigs were assigned to the BA group and seven pigs to the non-BA group. In the non-BA group, apnea was observed in four of seven cases, three of which resulted in death. None of the eight pigs in the BA group had respiratory arrest; notably, all survived. Hypotension was observed in some pigs in each group; however, there were no cases of bradycardia in either group. Statistical analysis showed that wearing BA significantly reduced the occurrence of respiratory and cardiac arrest (p=0.026) but not survival (p=0.077). No significant differences were found in other vital signs. Conclusions Wearing BA with adequate neck and chest protection reduced mortality and it was effective to reduce cardiac and respiratory arrest against shock wave exposure. Mortality from shock wave injury appears to be associated with respiratory arrest, and the avoidance of respiratory arrest may lead to survival.

Impact of Blue Light Intervention on Physiological Parameters and Circadian Rhythms: Insights from a Wistar Rat Model Study

Abstract Background: This study investigates the impact of blue light-emitting diode (LED) lighting on the physiological responses of Wistar rats. The circadian system plays a vital role in regulating physiological functions, allowing organisms to anticipate and adapt to environmental rhythms. The synchronization of endogenous circadian clocks with external cues, such as light-dark cycles, is essential for optimal performance. However, emerging technologies, like blue LED lighting, present new challenges to circadian rhythms, highlighting the need for continued research in this area. Methods: The study, approved by the Animal Ethics Committee, involved a total of 12 male Wistar rats, which were divided into two groups: a Control group and a blue light (BL) treated group, each consisting of six rats. The BL model was established by subjecting the rats to 12 h of BL exposure daily for a period of 3 months. Half of the rats were sacrificed at the end of this period, while the remaining rats were transitioned to normal light (NL) conditions for another 3 months before being euthanized. Blood and tissue samples were collected for analysis, including measurements of body weight, blood glucose, insulin, melatonin, lipid profile, as well as messenger RNA expression by reverse transcription-polymerase chain reaction and global DNA methylation by enzyme-linked immunosorbent assay. Results: The study found that BL exposure led to a higher monthly increase in body weight, elevated blood glucose levels, and decreased insulin and melatonin levels compared to controls. BL also caused a decrease in total cholesterol, triglycerides, high-density lipoprotein, and low-density lipoprotein. Gene expression analysis revealed downregulation of Period1, Cryptochrome1, and brain and muscle arnt-like protein 1 (Bmal1) in blood and hypothalamus, with significant upregulation upon returning to NL conditions. BL exposure induced hypermethylation in DNA sequences, which partially reverted after returning to NL conditions. Conclusion: Our study reveals the broad impact of BL/artificial light contributing to a higher incidence of obesity and metabolic disorders. With the rise of blue-rich LED lighting, more research is needed to understand its long-term effects on health. This study contributes to a deeper understanding of the intricate relationship between BL exposure and metabolic health, emphasizing the importance of informed decisions regarding lighting technologies to mitigate potential health risks.

Ameliorative effect of Curcuma longa against arsenic induced reproductive toxicity in Charles Foster rats

The present study aims to develop the antidote against arsenic induced male reproductive toxicity in animal models. The study was carried out on Charles Foster Rats after the approval from Institutional Animal Ethics Committee. A total of n=18, rats (12 weeks old) of an average weight of 160 ± 20 g were used for the study. The study group included n=6 control and n= 12 treated with sodium arsenite orally at the dose of 8mg/Kg b.w daily for 40 days. The n= 6 animals were dissected and rest n=6 was administered orally with Curcuma longa rhizome ethanolic extract at the dose of 600mg/Kg b.w per day for 40 days. At the end of the entire experiment, all the animals were dissected out and their reproductive organs were taken out, especially epididymis for sperm counts, sperm motility, sperm mortality, sperm morphology. The blood samples were collected for the hormonal assay (testosterone and luteinizing hormone), as well as for hematological and biochemical analysis. The study showed, high magnitude of degeneration in the reproductive organs of the rats in the arsenic treated group. There were degenerative fluctuations in the sperm counts, sperm motility, sperm mortality, sperm morphology and in the hormonal parameters, as well as in the hematological and biochemical parameters in the arsenic treated rats. But, after the administration of Curcuma longa, there was significant amelioration in all these parameters. Therefore, the present study shows that Curcuma longa plays vital role to combat arsenic induced male reproductive toxicity.

In Vivo Wound Healing Potential and Molecular Pathways of Amniotic Fluid and Moringa Olifera-Loaded Nanoclay Films.

Cutaneous wounds pose a significant health burden, affecting millions of individuals annually and placing strain on healthcare systems and society. Nanofilm biomaterials have emerged as promising interfaces between materials and biology, offering potential for various biomedical applications. To explore this potential, our study aimed to assess the wound healing efficacy of amniotic fluid and Moringa olifera-loaded nanoclay films by using in vivo models. Additionally, we investigated the antioxidant and antibacterial properties of these films. Using a burn wound healing model on rabbits, both infected and non-infected wounds were treated with the nanoclay films for a duration of twenty-one days on by following protocols approved by the Animal Ethics Committee. We evaluated wound contraction, proinflammatory mediators, and growth factors levels by analyzing blood samples. Histopathological changes and skin integrity were assessed through H&E staining. Statistical analysis was performed using SPSS software (version 2; Chicago, IL, USA) with significance set at p < 0.05. Our findings demonstrated a significant dose-dependent increase in wound contraction in the 2%, 4%, and 8% AMF-Me.mo treatment groups throughout the study (p < 0.001). Moreover, macroscopic analysis revealed comparable effects (p > 0.05) between the 8% AMF-Me.mo treatment group and the standard treatment. Histopathological examination confirmed the preservation of skin architecture and complete epidermal closure in both infected and non-infected wounds treated with AMF-Me.mo-loaded nanofilms. RT-PCR analysis revealed elevated concentrations of matrix metalloproteinases (MMPs) and vascular endothelial growth factor (VEGF), along with decreased levels of tumor necrosis factor-alpha (TNF-α) in AMF-Me.mo-loaded nanofilm treatment groups. Additionally, the antimicrobial activity of AMF-Me.mo-loaded nanofilms contributed to the decontamination of the wound site, positioning them as potential candidates for effective wound healing. However, further extensive clinical trials-based studies are necessary to confirm these findings.

Mutagenicity and safety evaluation of Ashwagandha (Withania somnifera) root aqueous extract in different models

Withania somnifera (Ashwagandha) also called as Indian ginseng, a revered herb from Indian traditional system of medicine is a rejuvenator and tonic (Rasayana) used for its varied benefits. The roots of ashwagandha exhibit properties like anti-inflammatory, aphrodisiac, anthelmintic, astringent, diuretic, stimulant and thermogenic. However, data of ashwagandha on its mutagenic effects are lacking. In the present study, in-vitro genotoxicity tests were used to evaluate the mutagenic potential of Ashwagandha Root Extract (ARE). Concentrations of 0.156 to 5.00 mg/plate ARE were used for conducting Bacterial reverse mutation test (BRMT). For chromosome aberration (CA) test ARE was used in concentrations of 0.25 to 2.00 mg/ml, and for micronucleus (MN) tests ARE concentrations of 500/1000/2000 mg/kg were used. Acute oral toxicity was conducted in Wistar rats (n = 25) as per the OECD guideline (#423) with doses of 500/1000/2000 mg/kg body weight in male Swiss albino mice for morbidity and mortality for 3 days. The BRMT and CA tests were conducted with and without metabolic activation (S9). The study was approved by the institutional ethics committee (IEC) and institutional animal ethics committee (IAEC). ARE failed to show any mutagenic effects up to a dose of 5 mg/plate in BRMT. Also, ARE did not show any clastogenic activity in doses up to 2 mg/ml in CA test and in micronucleus test up to 2000 mg/kg body weight. These results were observed with and without metabolic activation (S9) under the stated experimental conditions. No mortality, morbidity, or any clinical signs were observed up to 3 days following ARE administration. Ashwagandha root extract failed to show any mortality in doses up to 2000 mg/kg oral dosage and did not show any mutagenic (genotoxic) effects in high concentrations.

Evaluation of renal ischemia–reperfusion injury using CEUS in mice

BackgroundRenal ischemia–reperfusion injury (IRI) frequently occurs clinically. We investigated the value of contrast-enhanced ultrasonography (CEUS) in the evaluation of renal IRI levels in mice.MethodsThirty-six healthy adult male C57BL/6 mice (20–22 g) were randomly divided into the sham, 10 min, 20 min, 30 min, 40 min, and 50 min groups based on the time of renal warm ischemia by blocking the left renal pedicle, approved by the Institutional Animal Ethics Committee. Time-intensity curve (TIC)-derived parameters such as peak enhancement (PE) and wash-in perfusion index (WiPI) were produced using CEUS at 1 h and 24 h after IRI. The severity of kidney injury was detected by the renal tubular necrosis rate which was analyzed by hematoxylin and eosin staining at 24 h after IRI. The Spearman correlation coefficient was used to describe the correlations between PE and WiPI values and the renal tubular necrosis rate.ResultsThe PE and WiPI values decreased after IRI in the groups with a warm ischemia time ≥ 20 min. The renal tubular necrosis rate was significantly correlated with the PE value at 1 h (ρ = -0.802) and 24 h (ρ = -0.861) after IRI and the WiPI value at 1 h (ρ = -0.814) and 24 h (ρ = -0.853) after IRI (all p < 0.001).ConclusionTIC-derived parameters, including PE and WiPI values, can be used to evaluate the severity of renal IRI in mice. CEUS is a safe and effective technology for the detection of renal IRI.Relevance statementCEUS can evaluate the severity of renal ischemia–reperfusion injury by peak enhancement and wash-in perfusion index values selected from various time-intensity curve-derived parameters.Key points• Contrast-enhanced ultrasonography can evaluate the level of renal ischemia–reperfusion injury.• Peak enhancement and wash-in perfusion index are correlated with the renal tubular necrosis rate.• CEUS can detect changes in unilateral renal function without radiation.Graphical

The role of synbiotics in Alzheimer`s management

AbstractBackgroundQuercetin and Lactobacillus Brevis were used in the development of S‐SNEDDS for their antiinflammaging effect in Alzheimer`s diseaseMethodThe isotropic mixture was prepared by incorporating Capmul MCM, Tween 80 and Transcutol P as oil, surfactant and co surfactant phases respectively. Quercetin was then added. The probiotic mixture was selected as a solid carrier and drug loading was calculated. The droplet size and zeta potential was also determined. The formulation was also assessed for differential scanning colorimetry (DSC), powder x ray diffraction studies (PXRD), and field emission scanning electron microscopy studies (FESEM). Stability studies of the developed formulation was also carried out. Alzheimer’s disease was induced in rats by the use of aluminum chloride (100mg/kg) after taking necessary permission from the institution animal ethical committee. The Morris water maze (MWM) test was used to assess the animals 48 hours following the last treatment. Utilizing marketed ELISA kits, the concentrations of TNF, IL‐1, and IL‐6 were determined.ResultQuercetin was solubilized in the isotropic mixture. The drug loading was found to be 93.4±0.75% for the liquid SNEDDS (QUE‐L‐SNEDDS) formulation. The droplet size of QUE‐L‐SNEDDS was found to be 73.24±1.45nm and had a zeta potential of ‐23.21±1.34 mV. Whereas, the solid SNEDDS (QUE‐LB‐S‐SNEDDS) was found to be 74.56±1.23 nm and had a zeta potential of ‐21.43 mV, respectively. This indicated that there was no change in the L‐SNEDDS and S‐SNEDDS formulation. Further, the amorphous nature of quercetin was confirmed by the DSC, PXRD and FESEM studies. Aluminum chloride was successful in inducing Alzheimer`s disease in the rats. Impaired learning and memory impairments, increased short‐term memory loss, and decreased locomotor activity were all observed in the negative controls. The developed formulation was successful in reversing the inflammatory effects of Alzheimer’s disease.ConclusionThe present study reveals that Quercetin and Lactobacillus Brevis can be used as a symbiotic combination as well as an adjuvant therapy in the treatment of inflammation related to Alzheimer’s disease. The results reveal an interesting outcome at the preclinical level and warrants future studies aimed at other plant based polyphenolic compounds especially in neurodegenerative diseases such as Alzheimer’s disease.

Sequential ultrasound molecular imaging for noninvasive identification and assessment of non-alcoholic steatohepatitis in mouse models

Background and objectiveNoninvasive non-alcoholic steatohepatitis (NASH) assessment is a clinical challenge to the management of non-alcoholic fatty liver disease. We aim to develop diagnostic models based on sequential ultrasound molecular imaging (USMI) for the noninvasive identification of NASH in mouse models. MethodsAnimal experiments were approved by the Animal Ethics Committee of South China Agricultural University. Forty-nine C57BL/6 mice were divided into normal control, non-alcoholic fatty liver, NASH, and hepatitis groups. Sequential USMI was implemented using CD36-targeted microbubbles (MBs-CD36) and intercellular adhesion molecule-1 (ICAM-1)-targeted microbubbles (MBs-ICAM-1) to visualize hepatic steatosis and inflammation. The targeting signal of USMI was quantified as the normalized intensity difference (NID) with the destruction-replenishment method. Correlation analysis was conducted between the NID-MBs-CD36 and pathological steatosis score and between the NID-MBs-ICAM-1 and pathological inflammation score. Finally, diagnostic models combining NID-MBs-CD36 with NID-MBs-ICAM-1 were established for NASH diagnosis. ResultsMBs-CD36 and MBs-ICAM-1 were successfully prepared and used for sequential USMI in all mice. NID-MBs-CD36 values increased with the progression of steatosis, while NID-MBs-ICAM-1 values increased in parallel with the progression of inflammation. A strong positive correlation was identified between NID-MBs-CD36 and pathological steatosis grade (rs = 0.9078, P < 0.0001) and between NID-MBs-ICAM-1 and pathological inflammation grade (rs = 0.9071, P < 0.0001). Among various sequential USMI-based diagnostic models, the serial testing model showed high diagnostic performance in detecting NASH, with 95% sensitivity, 97% specificity, 95% positive predictive values, 97% negative predictive values, and 96% accuracy. ConclusionsSequential USMI using MBs-CD36 and MBs-ICAM-1 allows noninvasive grading of hepatic steatosis and inflammation. Sequential USMI-based diagnostic models hold great potential in the noninvasive identification of NASH.

ANTI-INFLAMMATORY ACTIVITY OF SIDDHA POLYHERBAL FORMULATION SEVVIYADHI CHOORANAM ON CARRAGEENAN INDUCED PAW EDEMA IN WISTAR ALBINO RATS

Objective: The aim of the study was to explore the anti-inflammatory activity of Siddha polyherbal formulation Sevviyadhi chooranam in Carrageenan induced paw edema in wistar albino rats, and compared with the standard drug Indomethacin. Methods: The Siddha polyherbal formulation Sevviyadhi chooranam indicated for sinusitis was prepared based on GMP (Good Clinical Practice) guidelines. Study procedure was approved by Institutional Animal Ethics Committee (IAEC). The experimental animals were measured for paw edema volume at 1, 2, 3, 4, 5 h using Plethysmometer (Model 7150 UGO Basile, Italy). Edema was expressed as mean increase in paw volume relative to control animals. And then, findings were compared with Indomethacin (Standard drug). Results: The findings revealed that test drug Sevviyadhi chooranam at higher dosage 200 mg/kg (Group V) had equal effect on anti-inflammatory activity with percentage protection of 93.2% when compared with the standard drug Indomethacin at about 40 mg/kg (Group III) with percentage protection 93.2%. However, the test drug Sevviyadhi chooranam at a higher dosage 200 mg/kg (Group V) with a percentage protection 93.2% was highly effective when compared with lower dosage about 100 mg/kg (Group IV) with a percentage protection 27.12%. Hence, the study resulted that the Siddha polyherbal formulation Sevviyadhi chooranam has an optimistic anti-inflammatory activity with more therapeutic value. Conclusion: The study concluded that the Siddha polyherbal formulation Sevviyadhi chooranam has a promising anti-inflammatory activity, probably due to the presence of biologically active phytocompounds. However, it is important to admit that there are some scientific evidences of the potential actions of these phytocompounds in anti-inflammatory activity.

Protective effect of Cordia myxa leaves extract in FCA-induced arthritis in rats

Background: Arthritis, an age-related chronic inflammatory autoimmune disease primarily characterized by joint pain, swelling, stiffness, cartilage destruction, is increasing tremendously at an alarming rate and also becoming a major risk factor for other health complications. Cordia myxa, family Boraginaceae is an herbal plant used since ancient times for its various significant pharmacological activities, including anti-inflammatory effects. Scientific studies reporting anti-arthritic potential of leaves extract of this plant are still lacking. Therefore, the present work has been designed to investigate the protective effect of ethanolic extract of Cordia myxa leaves in Freund’s complete adjuvant (FCA)-induced arthritis in rats. Methods: Ethanolic extract (100mg/Kg, 200mg/Kg and 400mg/Kg) was investigated for their efficacy in the management of FCA induced arthritis in rats. Various parameters like paw volume (using plethysmograph), paw thickness (using Vernier calliper), body weight, hematological and histopathological profile were observed. The protocol was approved by Institutional Animal Ethical Committee (MMCP/IAEC/94) and was according to CPCSEA guidelines. Results: The phytochemical screening reveals the presence of flavonoids, glycosides, alkaloids, saponins and carbohydrates in the ethanolic extract of Cordia myxa leaves. Promising results with significant improvement were discovered as evident from all determined parameters when compared with standard drug methotrexate, which might be due to the presence of flavonoids, glycosides, alkaloids in the ethanolic extract of Cordia myxa leaves. Conclusion: The present study supports the traditional use of Cordia myxa leaves as potential anti-arthritic agent which can be effective in Freund’s complete adjuvant (FCA)-induced arthritis in rats, however its mode of action is still need to be studied.

CONSUMPTION OF HYPERLIPIDEMIC DIET AND WATER INTAKE IN ALBINO RATS TREATED WITH ECLIPTA ALBA – ORIGINAL RESEARCH PAPER

Objective: A high-fat (HF) diet described as higher calories generating from fat instead of carbohydrate or protein. HF usually diets a typical low-carbohydrate diet. Methods: Thirty adult Albino rats of Wistar strain accurately weighing between 165 and 215 g utilized for the present the study. As per CPCSEA guidelines, the protocol was approved by the institutional animal ethical committee no. BMCH/IAEC/01 Anat/2015 dated June 4, 2015. Animals were maintained and fed with standard rat chow, HF diet, and clean drinking water ad libitum, commercially available and manufactured by the National Institute of Nutrition, Hyderabad. Results: Rate of food consumption increased in rats fed with the HF diet for 8 weeks and decreased significantly in rats treated with Eclipta alba for 1, 2, and 3 weeks progressively compared to control group. Water intake increased in the rats fed with an HF diet (43±2.1) compared to rats fed with a normal diet, decreased significantly in the rats treated with E. alba (31±1.4) compared to the HF diet-fed animals. Conclusion: The rate of food consumption increased in rats fed with the HF diet for 8 weeks and decreased significantly in rats treated with E. alba for 1, 2, and 3 weeks progressively compared to control group. Water intake increased in the rats fed with an HF diet compared to rats fed with a normal diet, decreased significantly in the rats treated with E. alba compared to the HF diet-fed animals

Guidelines for the establishment and functioning of Animal Ethics Commitees (Institutional Animal Care and Use Committees) in Africa.

Animals are used for scientific purposes across Africa to benefit humans, animals or the environment. Nonetheless, ethical and regulatory oversight remains limited in many parts of the continent. To strengthen this governance framework, the Pan-African Network for Laboratory Animal Science and Ethics brought together experts from 12 African countries to create an Africa-centric practical guide to facilitate the establishment and appropriate functioning of Institutional Animal Ethics Committees across Africa. The Guidelines are based on universal principles for the care and use of sentient animals for scientific purposes, with consideration of the cultural, religious, political and socio-economic diversity in Africa. They focus on 11 key elements, including responsibilities of institutions and of the Institutional Official; composition of the Committee; its responsibilities, functioning and authority; ethical application and review processes; oversight and monitoring of animal care and use and of training and competence; quality assurance; and the roles of other responsible parties. The intent is for African institutions to adopt and adapt the guidelines, aligning with existing national legislation and standards where relevant, thus ensuring incorporation into practice. More broadly, the Guidelines form an essential component of the growing discourse in Africa regarding moral considerations of, and appropriate standards for, the care and use of animals for scientific purposes. The increased establishment of appropriately functioning animal ethics committees and robust ethical review procedures across Africa will enhance research quality and culture, strengthen societal awareness of animals as sentient beings, improve animal well-being, bolster standards of animal care and use, and contribute to sustainable socio-economic development.

Nerve Injury and Peripheral Nerve Functional Loss From Injection and Suture Needles: An Experimental Study on Rats.

During invasive procedures involving needles, there is a chance of damage to peripheral nerves. Therefore, a method for experimentally simulating these situations is needed. In this study we aimed to evaluate peripheral nerve lesions caused by the entry and exit of an injection needle, nerve transfixion through a suture stitch, and the injection of saline solution and lidocaine into the nerve. After obtaining approval from the Animal Ethics Committee, we randomly divided 36 Wistar rats, weighing approximately 250 g each, into 6 groups (control, sham, suture, needle, saline, and lidocaine groups), with 6 animals in each group. All procedures were performed on the left paws of the rats. After the procedure, walking track analysis was performed to assess the walking function of rats for 8 weeks. Four months after the procedures, we performed bilateral electrophysiological studies (measuring the latency and amplitude of the electrical impulse in the peroneal nerve). Subsequently, the rats were euthanized, and bilateral tissue samples were collected from the peroneal nerve and tibialis cranialis muscle for morphometric histological analysis. In the walking track analysis, all groups showed normalization of walking functionality after 13 days. There was little histological change between the groups, and no functional loss related to the procedures was observed. Procedures involving the infusion of local anesthetic or saline solution, or suture needles, are safe with regard to peripheral nerve function loss. Accidents tend not to cause nerve impairment.