CO₂ inhalation is currently the most common method of euthanasia for laboratory rats and mice, and it is often used for further terminal blood sampling for clinical biochemical assays. Lately, this method has been criticized due to animal welfare issues associated with some processes that develop after CO₂ inhalation. The stress reaction and the value of the clinical laboratory parameters significantly depend on the used anesthetics, method, and the site of blood sampling. Especially in small rodents, an acute terminal state followed by a cascade of metabolic reactions that can affect the studied biochemical profile may develop and cause unnecessary suffering of animals. The aim of this study was to compare the stability of biochemical parameters of outbred Sprague Dawley rats and CD-1 mice serum collected after CO₂ inhalation or the intramuscular injection of tiletamine–zolazepam–xylazine (TZX). The serum content of total protein and albumin, cholesterol, triglycerides, aspartate aminotransferase (AST), alanine aminotr ansferase (ALT), alkaline phosphatase (ALP), total bilirubin, and creatinine was decreased by the injection of TZX in comparison with CO₂ inhalation. In addition, the levels of calcium, phosphates, chlorides and potassium were lowered by TZX vs. CO₂ administration, while the level of sodium increased. Finally, the level of the majority of serum clinical biochemical parameters in rats and mice tend to be overestimated after CO₂ inhalation, which may lead to masking the possible effect of anti-inflammatory drugs in animal tests. Injection anesthesia for small rodents with TZX is a more feasible method for terminal blood sampling, which also reduces the suffering of animals.

Introduction: The article presents the results of the functional tests to improve the assessment of MIA-induced osteo­arthritis development and the effectiveness of NSAID therapy. Materials and methods: In the study, 26 male SD rats were used. MIA-induced osteoarthritis was simulated in the right knee joint. After an intra-articular injection of MIA, the animals were treated with ibuprofen and meloxicam. Pain assessment was studied in the following functional tests: incapacitance (hind limb weight bearing) test, von Frey test (mechanical allodynia), grip strength test, and knee diameter measurement. At the end of the study, a histological analysis of the knee joint was performed. Results and discussion: An intra-articular MIA injection reduced 1.5 times the paw withdrawal threshold. In the rats that suffered MIA-induced osteoarthritis, the difference between the diameters of the intact and injected joints was 1.05 mm, compared to 0.03 mm difference in the control group. Hind limb weight bearing asymmetry was 89.5% when simulating MIA-induced osteoarthritis. The muscular hind limb grip strength in rats with MIA-induced osteoarthritis was significantly reduced on 3rd and 7th days after simulating osteoarthritis. Ibuprofen and meloxicam showed signif­icant efficacy in all the above tests, although ibuprofen effectiveness was more pronounced than that of meloxicam. Conclusion: The following functional tests were identified as the most significant and sufficient to assess the devel­opment of MIA-induced osteoarthritis and analgesic efficacy of NSAIDs: incapacitance test, allodynia test (von Frey filaments), measurement of hind limb grip strength and measurement of the diameter of the inflamed knee joint. The histological analysis made it possible to confirm the correspondence of the physiological response and pathological changes in the knee joint.

Diabetes mellitus is a serious threat to human health in both developed and developing countries. Optimal disease control requires the use of a diet and a combination of several medications, including oral hypoglycemic agents such as α-glucosidase inhibitors. Currently, the arsenal of available drugs is insufficient, which determines the relevance of studying new potent α-amylase inhibitors. We implemented the recombinant production of sea anemone derived α-amylase inhibitor magnificamide in Escherichia coli. Peptide was isolated by a combination of liquid chromatography techniques. Its folding and molecular weight was proved by ¹H NMR and mass spectrometry. The K_i value of magnificamide against human pancreatic α-amylase is 3.1 nM according to Morrison equation for tight binding inhibitors. Our study of the thermodynamic characteristics of binding of magnificamide to human salivary and pancreatic α-amylases by isothermal titration calorimetry showed the presence of different binding mechanisms with Kd equal to 0.11 µM and 0.1 nM, respectively. Experiments in mice with streptozotocin-induced diabetes mimicking diabetes mellitus type 1 were used to study the efficiency of magnificamide against postprandial hyperglycemia. It was found that at a dose of 0.005 mg kg^-1, magnificamide effectively blocks starch breakdown and prevents the development of postprandial hyperglycemia in T1D mice. Our results demonstrated the therapeutic potential of magnificamide for the control of postprandial hyperglycemia.

Many studies confirm that substances of natural origin have a pronounced affinity for type 2 diabetes mellitus (T2DM) therapeutic targets. At the moment, there is growing interest in bioactive peptides, phytochemicals, and drugs from other natural sources as highly effective, safe and promising antidiabetic agents. Natural sources are a promising resource for regulating several pathological pathways in T2DM. The review describes ways to mitigate insulin resistance and tissue sensitivity to glucose through PTP1β (protein tyrosine phosphatase 1β), GLP-1R (glucagon-like peptide receptor), DPP-4 (dipeptidyl peptidase-4), AMPK (adenosine monophosphate activated protein kinase), MAPK (mitogen-activated protein kinase). The review includes ways to regulate obesity and oxidative stress development through CCN3 (nephroblastoma overexpressed gene), PPAR-γ (peroxisome proliferator-activated receptor γ), Nrf2 (nuclear factor erythroid-related factor 2), FFAR (free fatty acid receptors), 11β-HSD1 (11β-hydroxysteroid dehydrogenase). Moreover, the review details the regulation of hyperglycemia through alpha-amylase inhibitors, as well as regulation of glucose metabolism through GFAT (glutamine fructose6-phosphate aminotransferase), FOXO1 (forkhead box protein O1), GLUT4 (glucose transporter type 4), PGC-1α (receptor gamma coactivator 1α activating peroxisome proliferator). The review examines the use of natural sources, from which low-molecular-weight and peptide compounds are used as T2DM targets modulators.