Double Administration Research Articles

ENCAPSULATION OF EUCALYPTUS LEAVES PHYTOPRODUCTS INTO LIPOSOMAL NANOPARTICLES AND STUDY OF THEIR ANTIBACTERIAL ACTIVITY AGAINST STAPHYLOCOCCUS AUREUS IN VIVO

Plant oils are high effective active pharmaceutical ingredients, but the extremely low bioavailability of lipophilic compounds limits their use in drugs. The encapsulation of essential oils into nanoparticles are actively studied. The aim of the research was to prepare liposomal forms of eucalyptus leaves extract and Chlorophyllipt and to study their antibacterial activity. Liposomal forms of Chlorophyllipt and eucalyptus oil extract was prepared using egg phosphatidylcholine and cholesterol by high pressure homogenization and sonication method. The average particle sizes of liposomal Chlorophyllipt and liposomal eucalyptus extract were 156.5 and 210.4 nm, respectively. Active pharmaceutical ingredient encapsulation in liposomal Chlorophyllipt and liposomal eucalyptus extract were at least 85% and at least 90%, respectively. Antibacterial activity of liposomal drugs was studied in white mice with model of staphylococcal infection, initiated by Staphylococcus aureus ATTC 209. The use of liposomal eucalyptus oil and liposomal Chlorophyllipt demonstrates no toxicity in mice. Both investigated drugs demonstrated antibacterial activity against Staphylococcus aureus in mice. A single administration of the liposomal drugs increased the survival rate up to 30-40% compared with model animals. Double administration of liposomal drugs both in liquid form and in lyophilized form increased the survival rate by at least 70 %. Moreover, in the group of animals treated liposomal eucalyptus oil, the number of surviving animals was 100%, while in a group of liposomal Chlorophyllipt it was 80%.

Ensuring veterinary well-being and productivity of pigs with an immunotropic drug

The purpose of this work is to realize the productive qualities of young pigs by increasing the effectiveness of specific prevention of infectious diseases. The object of the study was 45 pigs of the Landrace breed from 14 to 171 days of age, divided according to the principle of pairs of analogues into 3 groups of 15 heads. The animals of the 1st experimental group were injected with the immunotropic drug PigStim-V in a dose of 1.0 ml per head twice at the age of 14 and 21 days in order to stimulate nonspecific resistance of the body and increase the effectiveness of specific prevention of infectious diseases intramuscularly at the age of 14 and 21 days. Moreover, the second injection was carried out simultaneously with immunization against type 2 porcine circovirus infection. The tested immunotropic drug PigStim-V was administered to animals of the 2nd experimental group once, at a dose of 1.0 ml per head at the age of 21 days simultaneously with a commercial vaccine against porcine circovirus. Animals of the 3rd group served as biological control. The results of the study confirm the effectiveness of the use of the immunotropic drug PigStim-V as an adjuvant in the vaccination of pigs. Both single and double intramuscular injection of PigStim-V increases the titer of specific antibodies against porcine circovirus infection and against α, β and ε toxins of the microorganism Clostridium perfringens, prevents and increases the effectiveness of therapy of diseases of non-infectious etiology, stimulates growth. A more pronounced positive effect is observed with the double administration of an immunotropic drug.

Effects of double intravenous furosemide administration at different doses on clinical variables, electrocardiographic indices and serum electrolytes in miniature donkeys.

The intravenous (IV) usage of diuretics such as furosemide may cause changes in clinical variables, electrocardiographic (ECG) indices and some serum electrolytes concentrations in miniature donkeys. The purpose of this study was to evaluate the effects of double intravenous (IV) administration of furosemide at different doses on the clinical variables, ECG indices and serum electrolytes levels in miniature donkeys. A total of 21 clinically and para-clinically healthy miniature donkeys with a mean age of 6.00 ± 1.00 years and weight of 95.00 ± 5.00 kg were used. The animals were randomly divided into three groups of control (normal saline) and low- (1.00 mg kg-1) and high-dose (2.00 mg kg-1) of furosemide. Two IV injections with 12-hr interval were administered in each animal. The clinical and ECG parameters were measured at 0 (baseline), 2 (T2), 24 (T24) and 48 (T48) hr after drug administration. Serum concentrations of electrolytes were measured at T0, T24 and T48. The results of this study showed no changes in clinical variables (heart rate, rectal temperature and respiratory rate) and ECG indices (amplitude and duration of P, QRS and T waves and duration of PR, QT and RR intervals) as well as no significant changes in serum electrolytes (sodium, potassium, chloride, phosphorus, calcium and magnesium) levels. Although no significant effect on clinical variables, electrolytes levels and ECG indices after double IV administration of furosemide (1.00 and 2.00 mg kg-1) in miniature donkeys was observed, absence of these effects does not mean that furosemide had no pharmacological effect.

SJLA_2024_A2: Laparoscopic Colectomy with Lymph Node Navigation and Intracorporeal Anastomosis using Double Indocyanine Green Technique

Introduction: In laparoscopic colectomy for colorectal cancer, appropriate lymph node dissection and prevention of anastomotic leak are very important. Indocyanine green (ICG) images obtained through the laparoscopic procedure helped visualize lymphatic drainage vessels and inform decision-making to determine the vessels. Intracorporeal anastomosis has advantages such as earlier recovery of postoperative bowel function, shorter length of wound incision, reduction of intestinal mobilization range, and fewer incisional hernias. Background: We report the surgical technique and short-term results of 48 patients who underwent laparoscopic colectomy and intracorporeal anastomosis using the double ICG fluorescence technique from July 2020 to September 2023. Methods: Two injections of ICG (0.75 mg ×2) into the proximal and distal subserosa of the tumor preceded the surgical procedure after pneumoperitoneum. Intraoperative lymph node mapping by the Stryker1588, 1688AIM camera imaging system was visualized. Laparoscopic colectomy was performed according to the Complete Mesocolic Excision and Central Vascular Ligation concepts. Complete intracorporeal anastomosis was performed by a functional end-to-end or overlap or Delta anastomosis. After anastomosis, ICG (12.5 mg) was injected intravenously to check the intestinal blood perfusion at the anastomosis site. Results: Forty-eight patients (Sex; male: 23 and female: 25) (Tumor location; C: 20, A: 11, T: 8, D: 7, S:2) (pStage0:2, I:15, II: 16, III:10, and IV:5) were underwent. The median age was 74. median body mass index was 23. Median number of dissected lymph nodes was 18. The median operative time was 246 min. The median wound length was 3.5 cm. The median postoperative hospital stay was 7 days. Visualization of lymphatic flow was observable in 24 of 28 cases (85.7%). The transection line was changed in 2/48 cases (4.1%) after ICG fluorescence angiography. There were no intraoperative complications, and two patients had postoperative paralytic ileus. Discussion: We introduce a novel technique of double ICG administration during laparoscopic colon resection for colon cancer that enables improved lymphadenectomy and warrants the extent of intestinal resection. It contains the intrinsic solution to the potential risks related to more intense dissection for extended lymphadenectomy. This is an immediate assessment of blood supply to the anastomosed intestinal walls. ICG fluorescence has immanent advantages as a result of its very low toxicity, high sensitivity, fast feedback, and absence of radiation. It is helpful to determine the extent of a bowel resection and can avoid leaving positive lymph nodes behind. It was revealed that ICG accurately evaluates the blood supply to anastomotic sites in real time and can detect organ ischemia before reconstruction is performed. This is particularly useful in laparoscopic surgery, as detection of impaired blood supply is difficult and even more for intracorporeal anastomosis. Lower anastomotic leak and reoperation rates were revealed with the use of ICG in several studies. Lymph nodes’ metastases are the most important prognostic factor in colon cancer, which has a poor prognosis and high recurrence. The use of ICG in the identification of lymph flow and lymphadenectomy basin in colon cancer patients has been previously reported. Lymph drainage of the transverse colon is complicated, and understanding this lymphatic drainage system is difficult, making ICG fluorescence particularly useful in right-sided and splenic flexure colon cancer patients. Conclusion: ICG fluorescence lymphangiography in laparoscopic colectomy for colon cancer allows visualization of lymphatic flow and may complement more reliable lymph node dissection. In addition, the evaluation of intestinal perfusion using ICG contributes to the reduction of anastomotic leakage and enables safer intracorporeal anastomosis.

Assessment of the safety and clinical efficacy of the Brainmax in therapy of non-demented patients with a mild cognitive impairment

The assessment of the clinical efficiency and safety of the drug Brainmax and its influence on the degree of functional recovery in the treatment of patients with non-dementia cognitive disorders with this drug. An open multicenter randomized study included 60 patients of 18-55 years with light and moderate CI, having complaints of the cognitive spectrum. They used a clinical and neurological study using generally accepted scales and tests (MoCA, MMSE, MFI-20 tests, Schulta, DSST tests and an assessment of the quality of life of SF-36). Patients were randomized in two groups comparable by age and gender. Group 1 was treated with Brainmax per os twice every day for 14 days. After 10-days rest they received same medication for another 14 days. Group 2 was treated with Brainmax per os twice every day for 14 days, without the continuation. The total duration of the study was 40 days, the assessment of their condition was carried out on the 1st day (visit 1), after 15 days (visit 2) and after 40 days (visit 3) using the indicators of the above scales and tests in comparison with the background data. Safety assessment was carried out by the presence and structure of undesirable phenomena. The use of Brainmax led to a significant improvement in cognitive performance according to all generally accepted scales and tests (concentration and maintaining of attention, working memory, visual-constructive skills, volume and speed of attention speed, information processing and executive functions), as well as to the decrease severity of asthenia and improvement of the quality of life. Brainmax has shown a good safety profile, tolerability and clinical efficacy in the treatment of young and middle-aged patients with non-demented cognitive impairment. Significant improvement was observed both with single and double course administration of the drug, but a significantly better effect was noted after its repeated course, which reflects, among other things, the cumulative effect of the active substances of this drug and makes longer use of the drug Brainmax justified and appropriate in these categories of patients. The data obtained allow us to recommend the wider use of the drug Brainmax in clinical practice for the treatment of CI in patients of different ages, which will optimize therapy and improve the course and outcome of the disease.

Effects of human lactoferrin under conditions of neurotoxic exposure: experimental research

BACKGROUND: Translational research using laboratory animals aimed at revealing the features of the pathogenesis of Parkinsons disease serve as a tool for finding new therapeutic strategies.
 AIM: Was to investigate the effects of human lactoferrin (a multifunctional globular glycoprotein) on behavior the 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine-treated mice as the model of dopaminergic neurons loss.
 MATERIALS AND METHODS: Nigrostriatal dopaminergic injury was induced by single administration of 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (40 mg/kg) to five-month-old C57Bl/6 mice. Behavioral functions were assessed in the open field and rotarod tests and by the stride length analysis.
 RESULTS: Preliminary administration of lactoferrin resulted in a significant reduction in the severity of nervous system lesions induced by 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine. The positive effect of lactoferrin on the exploratory behavior of animals disturbed by neurotoxin, depending on the time of administration, was revealed. Exogenous protein with double preliminary administration had a protective effect on the change in body weight of mice after acute 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine exposure. This suggests a reduction in systemic toxic effects against the background of lactoferrin therapy.
 CONCLUSION: The results obtained indicate the possibility of the potential use of lactoferrin as a promising therapeutic agent in the treatment of neurodegenerative diseases.

Evaluation of the Effects of GnRH and hCG Administrations during and after Insemination on Fertility Parameters of Dairy Cows

Background: Getting pregnant and maintenance of the pregnancy in cows is crucial for the dairy farmers for sustainable production. Many methods are used for this purpose, one of them is the administration of gonadotropins at insemination and later in the early embryonic period. In this study, it was aimed to investigate the effects of different gonadotropin applications on fertility. Methods: A total of 406 dairy cows were divided randomly into five treatment groups according to GnRH (10 µg buserelin acetate, im) and hCG (1500 IU, im) administrations in different time periods at AI and post AI as; GI (n=84, d7; hCG), GII (n=106, d7; GnRH), GIII (n=62, d0 and d7; hCG), GIV (n=80, d0 and d7; GnRH), GV (n=74, d0 and d7; placebo: 2.5 ml 0.9% NaCl). Pregnancy control was done on day 30 and 45 post AI. Fertility parameters (conception rate, first service pregnancy rate, pregnancy per AI, embryonic loss) were calculated with the obtained data. Result: Our findings indicated that double dose hCG administration caused an improvement on the fertility parameters in cows with greater than 150 DIM. Also, conception rates of the cows with higher parity, milk yield and lower BCS improved with the administration of hCG on d0 and d7. We concluded that double-dose hCG administrations on d0 and d7 can be used to improve fertility after 150 DIM in cows with higher parity and milk yield.

Dynamics of cell-free DNA levels in the in vivo LPS-induced inflammation model

An increased concentration of extracellular cell free DNA (cfDNA) is a distinctive characteristic of pathologies that mainly occur in acute inflammation (myocardial infarction, sepsis, stroke, trauma). The increase of cfDNA in chronic inflammatory processes, oncological, autoimmune diseases is less significant and is mainly due to aberrant cell death processes. One of such diseases is systemic lupus erythematosus (SLE). It has recently been shown that, in addition to increased cfDNA concentration, the degree of inflammation can reflect the N/L index (neutrophil to lymphocyte ratio), being a simple and informative marker of disease activity in patients with SLE. The aim of the study was to study the dynamics of the level of cfDNA and the N/L index in the model of LPS-induced inflammatory response as observed in intact mice, and their relation to the phenotypic heterogeneity of model SLE. We used female hybrid mice (C57Bl/6xDBA/2) F1 and female DBA/2 mice at the age of 6-8 weeks. LPS of E. coli strain 111: B4 (Sigma) was injected intraperitoneally once at doses of 10 ng, 1 g and 100 g per mouse in PBS. The control group was injected with the appropriate volume of buffer. The TNF-binding domain of the variola virus CRMB protein was used as an inhibitor of TNF, which was administered 30 min before the introduction of LPS. The dynamics of the response to LPS was assessed after 4, 8, 11, 24 hours by the N/L index and the level of cfDNA; at the zero point, the parameters were determined before the introduction of LPS. A day after a single injection of LPS at a dose of 1 g/mouse, a SLE model was induced on the same hybrid mice (double intravenous administration with an interval of 6 days of spleen cells of the DBA/2 line, 60-70 106 cells each). Three months later, with proteinuria of 3 mg/ mL or more, mice were assigned to the SLEnephritis+ group, with a protein of less than 3 mg/mL, to the SLEnephritis- group. Statistical processing of the results was carried out by nonparametric statistics using the MannWhitney test. Differences were considered statistically significant at p 0.05. It was found that the change in the N/L index, as well as the change in the level of cfDNA, depends on the dose of LPS administered. It was shown that the level of cfDNA reaches its maximum after 8 and 11 hours after the introduction of LPS is reliably reduced when using the inhibitor TNF. A retrospective analysis indicates that there is a definite relationship between the response of intact mice to LPS before induction of cGVHD, and their subsequent division into variants of SLEnephritis + and SLEnephritis - in the course of disease development.

Efficacy of botulinum toxin in a patient with neurogenic detrusor overactivity

Neurogenic detrusor overactivity is a dangerin terms of the formation of vesicoureteralreflux, pyelonephritis, and chronic kidney disease. The standard treatment for neurogenic detrusor overactivity is intermittent catheterization of the bladder in combination with M-cholinoblockers, which is not always sufficient. In the article, the authors briefly outlined the physiological basis of intradetrusor injections of botulinum toxin in neurogenic detrusor overactivity in patients with myelodysplasia who underwent surgery for spinal hernias, tumors of the spinal canal, as well as patients with spinal trauma. The technology of intradetrusor injections of botulinum toxin is shown, possible complications are described. The effectiveness of intradetrusor injections of botulinum toxin is shown by a specific clinical example — a description of the medical history of a child who has a history of surgical treatment for a spinal hernia in the newborn period. After the operation, neurogenic detrusor overactivity was revealed, insensitive to treatment with M-cholinoblockers. Complications were vesicoureteral reflux and recurrent pyelonephritis. The results of cystometry before and after double administration of botulinum toxin are shown. Detrusor hypertension was stopped, pyelonephritis remission was achieved, trabecularity of the bladder mucosa was stopped. Simultaneously with the introduction of botulinum toxin, endoscopic correction of vesicoureteral reflux was performed.

Development of a Method and a Scheme for the Use of a Live Recombinant Vaccine "GamLPV". Safety and Tolerability of Double Intranasal Vaccination of Healthy Adult Volunteers

Introduction. The increase in the incidence of whooping cough in children and adults of different age groups justifies the need for their revaccination and the development of new, acceptable for these purposes. This work is devoted to substantiating the design of a clinical trial and describing the results of a comparative study of the safety and tolerability of the drug "GamLPV" with two-fold intranasal administration to healthy adult volunteers using two methods. The choice of the scheme and method of administration of the drug is justified. The serological structure of the population of adults aged 18–40 years living in Moscow and the Moscow region is characterized.Aim. Determination of the safety and tolerability of the drug with a double intranasal administration of the vaccine by drip method and spraying through an actuator.Materials and methods. A randomized placebo-controlled trial included 50 healthy male and female volunteers aged 18 to 40 years who met the inclusion criteria. The volunteers were divided into 2 groups of 25 people: a drip method of administration and spraying through an "actuator". By both methods, the drug was administered twice with an interval of 60 days.Results and discussion. Serological analysis of the population of healthy volunteers at the prescreening stage justified the inclusion in the study of volunteers containing anti-pertussis antibodies in the blood (IgG ≤40 Ed/ml). A comparison of the results of preclinical studies on an experimental model of non-human monkeys and the first phase of a clinical study of GamLPV allowed us to propose two methods of double administration of the drug as a promising vaccination scheme for volunteers. A comparative randomized study shows the safety of using the proposed scheme for vaccination of adult volunteers.Conclusions. Both proposed methods of double administration can be used to plan a multicenter study to research the immunogenicity and protective activity of GamLPV.

THE INFLUENCE OF PHOTOSENSITIVE PEPTIDOMIMETICS ON WEIGHT INDICES OF IMMUNE ORGANS OF EXPERIMENTAL ANIMALS WITH TRANSPLANTABLE LEWIS LUNG CARCINOMA

The use of photodynamic therapy of drugs capable of selective accumulation in the tumor or in affected cells, in particular photosensitive peptidomimetics, increases its effectiveness in various treatment schemes. Aim. Determination of weight indices (WI) of thymus and spleen in animals with carcinoma after photodynamic therapy with peptidomimetics: LMB002 and LMB033. Methods. Studies of WI of immune organs were carried out in mice of the C57 Black line on the 28th day after transplantation of Lewis lung carcinoma. The therapeutic effect was studied using photosensitive peptidomimetics: LMB002 and LMB033. The natural antibiotic gramicidin C was used as a control. Results. Comparison of the WI of the spleen of intact animals and animals with tumors showed a twofold (p< 0.05) increase in animals with tumors. As for the WI of the thymus, a tendency to its increase was observed in animals with tumors, compared to intact ones, but no significant difference was found. The following results were obtained: a decrease in the WI of the spleen in animals with tumors and the introduction of placebo and an increase in the WI of the spleen in animals treated with gramicidin C; an increase in spleen WI and a slight decrease in thymus WI under the influence of LMB002, an increase in spleen WI under the influence of LMB033, while the thymus WI did not change. Conclusions. It was established that the WI of the spleen during therapy with photosensitive peptidomimetics LMB033 and LMB002 increased in all groups of experimental animals compared to intact ones. The most significant effect on the spleen index was observed for the schemes of double administration of LMB033 and double phototherapy. No significant changes in the weight index of the thymus during therapy with photosensitive peptidomimetics LMB033 and LMB002 were found.

Effect of a combination of altrenogest and double PGF2α administrations on farrowing variation, piglet performance and colostrum IgG

The variation of gestation length in sows leads to difficulties performing farrowing supervision. The present study was performed to investigate whether oral administration of altrenogest until 112 days of gestation and double administration of PGF2α at 113 days of gestation can synchronise the onset of parturition in sows and minimise deleterious effects on the incidence of stillbirths and colostrum quality. Additionally, the effects of synchronised farrowing on colostrum yield and piglet birth weight, colostrum intake and survival rate of piglets until seven days of postnatal life were also investigated. In total, 193 Landrace x Yorkshire crossbred sows were randomly allocated according to parity number into two groups, i.e. control (n = 95) and treatment (n = 98). The control sows were allowed to farrow naturally. The treatment sows were orally administered 20 mg per day of altrenogest for four days from 109 to 112 days of gestation and were administered PGF2α twice on day 113 of gestation. Individual body weight at birth and 24 h after birth of piglets in all litters were determined in both control (n = 1609) and treatment (n = 1707) groups. Colostrum consumption of all piglets, colostrum yield, colostrum IgG and serum progesterone of sows were determined. On average, the total number of piglets born per litter were 17.0 ± 3.1. The proportion of sows farrowed before 114 days of gestation was higher in the control than the treatment group (8.4% and 2.0%, respectively, P = 0.05) and 92.8% of sows in the treatment group farrowed on day 114 of gestation. The percentage of stillborn piglets per litter did not differ significantly between control and treatment groups (4.5% and 4.6%, respectively). Colostrum yield of sows did not differ between control and treatment groups (5.52 ± 0.13 and 5.28 ± 0.12 kg, respectively, P = 0.174). However, colostrum intake of piglets was lower in the treatment than the control group (354.7 ± 6.6 and 381.2 ± 7.0 g, respectively, P = 0.012). Colostrum IgG was higher in the control than the treatment group (41.2 ± 1.1 and 37.3 mg per ml, P = 0.013). In conclusion, altrenogest treatment from 109 to 112 days and double administrations of PGF2α on day 113 of gestation can control gestation length in sows. No deleterious effects of this protocol on the incidence of stillbirths and sow colostrum yield were detected. However, piglet colostrum intake and colostrum IgG were compromised. Thus, care of newborn piglets in the treatment group should be considered.

Evaluation of the genotoxicity of the food dye tartrazine in a micronucleus test in vivo

Introduction. Food azo dye Tartrazine (E102) is widely used in the production of food, pharmacological and cosmetic products. Despite the approval for use, interest in a comprehensive assessment of the impact of food colours, especially synthetic ones, on health continues unabated. The analysis of literature evaluating the genotoxicity of Tartrazine in vivo studies revealed some inconsistencies in the results, that showed the possibility to test a retail food colouring in one of recommended tests. Materials and methods. The Tartrazine genotoxicity (produced in India, purity 88.37%) was studied in the micronucleus test on male mouse bone marrow cells (hybrids F1 CBA x C57Bl6/j). The test substance was double enteral administrated in the dose range 250-2000 mg/kg. The frequency of polychromatophilic erythrocytes (PCEs) with micronuclei (MNs) was estimated by the analysis of 4000 PCEs. The proportion of PCE among all erythrocytes was determined by analyzing 1000 cells per animal. Results. There was no increase in the frequency of micronucleated polychromatophilic erythrocytes over a concurrent negative control with double enteral administration of Tartrazine in all doses studied. The proportion of PCEs among all erythrocytes did not change. Limitations of the study are due to the methodology of the test: only cytogenetic disorders in a single tissue were analyzed under conditions of double enteral administration of the studied sample. Conclusion. The sample of the food dye Tartrazine (E 102) in the dose range of 250-2000 mg/kg did not show cytogenetic activity in the in vivo micronucleus test on mouse bone marrow cells after a double enteral administration.

Double administration of self-complementary AAV9NDUFS4 prevents Leigh disease in Ndufs4-/- mice.

Leigh disease, or subacute necrotizing encephalomyelopathy, a genetically heterogeneous condition consistently characterized by defective mitochondrial bioenergetics, is the most common oxidative-phosphorylation related disease in infancy. Both neurological signs and pathological lesions of Leigh disease are mimicked by the ablation of the mouse mitochondrial respiratory chain subunit Ndufs4−/−, which is part of, and crucial for, normal Complex I activity and assembly, particularly in the brains of both children and mice. We previously conveyed the human NDUFS4 gene to the mouse brain using either single-stranded adeno-associated viral 9 recombinant vectors or the PHP.B adeno-associated viral vector. Both these approaches significantly prolonged the lifespan of the Ndufs4−/− mouse model but the extension of the survival was limited to a few weeks by the former approach, whereas the latter was applicable to a limited number of mouse strains, but not to primates. Here, we exploited the recent development of new, self-complementary adeno-associated viral 9 vectors, in which the transcription rate of the recombinant gene is markedly increased compared with the single-stranded adeno-associated viral 9 and can be applied to all mammals, including humans. Either single intra-vascular or double intra-vascular and intra-cerebro-ventricular injections were performed at post-natal Day 1. The first strategy ubiquitously conveyed the human NDUFS4 gene product in Ndufs4−/− mice, doubling the lifespan from 45 to ≈100 days after birth, when the mice developed rapidly progressive neurological failure. However, the double, contemporary intra-vascular and intra-cerebroventricular administration of self-complementary-adeno-associated viral NDUFS4 prolonged healthy lifespan up to 9 months of age. These mice were well and active at euthanization, at 6, 7, 8 and 9 months of age, to investigate the brain and other organs post-mortem. Robust expression of hNDUFS4 was detected in different cerebral areas preserving normal morphology and restoring Complex I activity and assembly. Our results warrant further investigation on the translatability of self-complementary-adeno-associated viral 9 NDUFS4-based therapy in the prodromal phase of the disease in mice and eventually humans.

Pivotal role of micro-CT technology in setting up an optimized lung fibrosis mouse model for drug screening

Idiopathic pulmonary fibrosis (IPF) is a progressive disease with no curative pharmacological treatment. The most used animal model of IPF for anti-fibrotic drug screening is bleomycin (BLM)-induced lung fibrosis. However, several issues have been reported: the balance among disease resolution, an appropriate time window for therapeutic intervention and animal welfare remain critical aspects yet to be fully elucidated. In this study, C57Bl/6 male mice were treated with BLM via oropharyngeal aspiration (OA) following either double or triple administration. The fibrosis progression was longitudinally assessed by micro-CT every 7 days for 4 weeks after BLM administration. Quantitative micro-CT measurements highlighted that triple BLM administration was the ideal dose regimen to provoke sustained lung fibrosis up to 28 days. These results were corroborated with lung histology and Bronchoalveolar Lavage Fluid cells. We have developed a mouse model with prolonged lung fibrosis enabling three weeks of a curative therapeutic window for the screening of putative anti-fibrotic drugs. Moreover, we have demonstrated the pivotal role of longitudinal micro-CT imaging in reducing the number of animals required per experiment in which each animal can be its own control. This approach permits a valuable decrease in costs and time to develop disease animal models.

Abstract 218: Novel SOD mimetics GC4419 & GC4711 induce synergistic tumoricidal effects combined with radiotherapy in lung cancer models

Abstract Introduction: Superoxide dismutase mimetics (SODm) synergize with radiation (IR) by converting radiolytically produced superoxide to hydrogen peroxide. Cancer cells lacking catalase function fail to clear H2O2 resulting in increased tumoricidal load. In addition, complex interactions through redox signaling and immune interplay are suggested mechanisms of action. Our pre-clinical studies with the SODm GC4419 (AVA) indicated that increased inflammatory, TNFα, and apoptosis signaling contribute to treatment synergy that was most effective at hypofractionation (Sci Transl Med. 13:593). The analogue GC4711 (GC) is currently in clinical trials for SBRT against NSCLC and pancreatic cancer. However, optimal combination schedules and molecular mechanisms are largely unknown. Here, we present optimized combination schedules using human lung tumor xenografts regarding drug administration timing and dosage, as well as local and abscopal tumor control in a syngeneic lung tumor model. Further, we tested various cell stress and immune pathways to identify targets of synergistic IR-drug interaction. Methods: Female athymic nude and C57BL/6 mice were inoculated with H1299 and LLC, respectively. H1299 tumors were irradiated with 10-15 Gy and GC was given once or twice daily (24 or 2x20 mg/kg, respectively) over 5 days, including untreated controls. LLC tumors were set up as an abscopal model and mice received combination treatment with GC and 15 Gy, 15 Gy only, or GC only, including untreated controls. In vitro, lung cancer cell lines were pre-treated with AVA or GC 30 min prior to IR and assayed for various immune or DNA damage-related endpoints. Results: The anti-tumor efficacy of single or double GC administration showed nonlinear dose-dependence indicating complex drug-IR interplay. Optimal drug administration appears to be around 3 hours prior to irradiation while time-of-day of treatment has no observable effect in this model. In the LLC double tumor model, GC in combination with IR induced significant and pronounced abscopal tumor control. In vitro, analysis revealed GC action related to e.g. 53BP1, yH2AX, Akt, R-loop formation and replication stress, and interaction with various immune pathways. Discussion and Conclusions: The radiotherapeutic effect can be vastly increased with superoxide dismutase mimetics, both within and outside the treatment field. Combination therapy requires screening of schedules regarding timing, drug dosage, and IR dose and fractionation to achieve optimal tumoricidal effects. SODm synergize with IR acting on various molecular targets that can be harnessed to overcome treatment resistance. Secondary tumor control is a promising new venue for IR-SODm combination therapy. Moreover, redox-immune interaction opens new therapeutic applications with combined immune checkpoint inhibitors for unprecedented tumor control and cure rate. Citation Format: Britta Langen, Laurentiu Pop, Zengfu Shang, Moito Iijima, James Nicholson, Mingming Yang, Michael D. Story. Novel SOD mimetics GC4419 & GC4711 induce synergistic tumoricidal effects combined with radiotherapy in lung cancer models [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2022; 2022 Apr 8-13. Philadelphia (PA): AACR; Cancer Res 2022;82(12_Suppl):Abstract nr 218.

Investigations on dynamics of juvenile hormone mimics on the economic parameters of silkworm, Bombyx mori L.

Juvenile hormone mimics are known to prolong larval life in insects and thus in sericulture, it has been shown to boost good cocoon and silk yield. In the present experiment, the double hybrid (FC1XFC2) silkworm was utilized and analysed the performance of economic traits. The influence of juvenile hormone mimics ie., phytojuvenoids from Pinus (Pinus roxburghii), Custard apple (Annona squamosa) and Tapioca (Manihot esculenta), a commercially available juvenile hormone mimic Serimore and Pyriproxyfen has been recorded as quantitative characters of silkworm when administered topically with two different doses at two distinct larval instars. The quantum of expression of economic trait like cocoon weight, shell weight, shell ratio, filament length and denier were found to be increasing with the increase in the number of larval treatments which is double administration at the rate of 20% and 30% concentration for Pinus and Tapioca treated larvae. In case of custard apple treated larvae, double treatment at 20% concentration showed best results. Single treatment at 5µl and 10µl concentration of pyriproxyfen gave similar outcome with respect to all economic parameters. Among all, Pinus was found to be on par with Serimore for all the economic characters.

Influence of hormonal synchronization protocols in heifers-recipients on their pregnancy rate with the use of frozen-thawed embryos

At the moment, there are a great number of protocols enabling to prepare recipients for embryo transfer. However, because of their effectiveness variety on cattle breeding farms the issue of searching of practical schemes that allow successful reproductive cycle synchronization in recipients with the achievement of their highest pregnancy rate becomes relevant. Based on the information above, the aim of the study was to determine the most effective estrous cycle synchronization protocol in recipient heifers before embryo transfer which provide the high level of their pregnancy rate with the use of frozen-thawed embryos. The research was carried out on the beef cattle in Samara region of Russia. In the course of the study, the heifers (n=107) were selected as the potential recipients and then divided into three groups depending on the applied protocol for the estrous cycle synchronization. All the recipients were transferred with frozen-thawed embryos obtained in vivo from Aberdeen Angus donor cows. The doses of the drug products indicated below were administrated per one animal. During the study, it was found that in group I where we used a protocol included a double administration of 2 ml of the drug "Oestrophan" (Czech Republic) with an interval of 11 days estrous cycles were synchronized in 72.73±0.80 percent of animals. The pregnancy rate in group I was 29.17±0.97 percent. In group II where we applied a protocol included a double administration of "Surfagon" (Russia) at a dose of 10 ml and Oestrophan with an interval of 11 days estrous cycles were synchronized in 50.0±0.86 percent of animals. The pregnancy rate in that group was 61.11±1.22 percent. In group III that included the application of intravaginal progesterone implant CIDR with a simultaneous administration of Surfagon at a dose of 2 ml and at the time of CIDR extraction the administration of 2 ml of Oestrophan, estrous cycles were synchronized in 63.16±0.80 percent of animals. The pregnancy rate in that group was 66.67±1.00 percent. Thus, the group III was characterized by the highest pregnancy rate that was achieved by the application of the protocol included intravaginal progesterone implant CIDR in combination with both medical products Surfagon and Oestrophan.

Morphological changes in the liver of experimental animals in the early stages after correction of exposure to high doses of paracetamol

Introduction. Acetaminophen exhibits hepatotoxicity in overdose, which is the leading cause of drug-induced liver failure. The aim of the research was to study early morphological changes in the liver of experimental animals under the influence of high doses of paracetamol and after correction with hepatoprotectors. Materials and methods. The corrective effect of various drugs - ademetionine, ethylmethylhydroxypyridine succinate and oxymethyluracil after exposure to high doses of acetaminophen was studied. Acetaminophen was administered to experimental rats once intragastrically at a dose of 1 g/kg, animals received hepatoprotectors twice (after 1 and 24 hours) or four times (after 1, 24, 48, 72 hours) after the toxicant. Morphological studies of the liver of experimental animals were carried out. Results. Pathomorphological changes were revealed on rat liver preparations after exposure to paracetamol, the most pronounced after 72 hours of exposure. Oxymethyluracil has a protective effect on the structure of the liver during acute exposure to high doses of paracetamol, comparable to the well-known drugs “Heptor” and “Mexidol”. In addition, the protective effect of oxymethyluracil appears earlier. Limitations. Morphological studies of the liver of 90 white male rats were carried out for studying the corrective action of hepatoprotectors after acute exposure to high doses of acetaminophen and after double (after 1 and 24 hours) or quadruple (after 1, 24, 48, 72 hours) administration of ademethionine, ethylmethylhydroxypyridine succinate and oxymethyluracil, which allowed to determine an effective means to reduce the toxic effect of paracetamol on the liver. Conclusion. The data obtained may have a certain practical value in terms of the prospective use of oxymethyluracil as an additional effective agent for reducing the toxic effect of paracetamol.

Control of parturition in hyperprolific sows by using altrenogest and double administrations of PGF2α

The aim of the present study was to develop a protocol to reduce the variation in gestation length and synchronise the onset of parturition in sows by using altrenogest in combination with double administrations of prostaglandin F2alpha (PGF2α). In total, 188 Landrace x Yorkshire crossbred sows with parity numbers 3.1 ± 1.6 were included in the experiment. The sows were classified into two groups: CONTROL (n = 94) and TREATMENT (n = 94). CONTROL sows were allowed to farrow naturally, and TREATMENT sows were orally administered 20 mg/day of altrenogest starting when they entered the farrowing house (107.0 ± 2.0 days) until 113 (TREAT–113, n = 18), 114 (TREAT–114, n = 29) and 115 (TREAT–115, n = 47) days of gestation. The altrenogest-treated sows were administered PGF2α twice 6 h apart at 24 h after the withdrawal of altrenogest. The litters were randomly selected (25 and 26 litters from CONTROL and TREATMENT groups, respectively) to determine individual body weight at birth and at 24 h after birth. Gestation length of sows that farrowed naturally averaged 115.1 days (range 111–118), whereas gestation length of altrenogest-treated sows averaged 115.1–116.3 days (range 114–118). The colostrum yield of sows averaged 4.25 ± 1.19 kg and was not affected by the treatment (P > 0.05). Colostrum IgG in the CONTROL group was higher than in the TREAT–114 and TREAT–115 groups (P < 0.05) but did not differ significantly compared to the TREAT–113 group. The proportion of sows that farrowed during working hours (0700–1700 h) in the TREAT–113 group (72.3%) tended to be higher than in the CONTROL (46.4%, P = 0.053). The interval from the last altrenogest treatment until farrowing in the TREAT–113 group was longer than in the TREAT–114 and TREAT–115 groups (62.8, 40.7 and 34.6 h, respectively, P < 0.05). Similarly, the intervals from the first PGF2α administration to the onset of parturition in the TREAT–113 group (38.8 ± 3.8 h) was longer than TREAT–114 (21.9 ± 3.5 h, P = 0.002) and TREAT–115 (25.5 ± 3.7 h, P = 0.016) groups. However, the incidence of stillbirths in the TREAT–113, TREAT–114 and TREAT–115 groups was higher than in the CONTROL (16.4, 17.2, 11.8 and 5.8%, respectively, P < 0.05). In conclusion, altrenogest supplementation in combination with double administrations of PGF2α can reduce the variation in gestation length and synchronise the onset of parturition in sows. However, its side effects on the incidence of stillbirths should be considered.