Anabolic steroids induced changes at the level of protein expression: Effects of prolonged administration of testosterone and nandrolone to pigs.

Synthetic derivatives of steroid hormones, specifically anabolic-androgenic steroids (AAS), have gained prominence due to their observed benefits in enhancing meat quality. The study replicated the administration of banned AAS and investigated their impacts on pigs to contribute to the understanding of animal biochemistry and to explore the feasibility of detecting AAS administration by employing a non-targeted analysis. The effects were corroborated by evaluating changes in the expression of selected proteins, as well as examining haematological and biochemical profiles and histological alterations. Exposure to AAS influenced the expression of proteins related to drug-metabolizing enzymes, muscle and lipid metabolism, kidney function, reproductive processes, immune system functions, and carcinogenic changes. The effects of AAS appear intricate and contingent on factors such as the specific drug used, dosage, and duration of administration. The results underscore that protein expression analysis holds promise as a valuable tool for detecting illicit AAS use in the fattening process.

The Superior Effect of Radiofrequency With Targeted Ultrasound for Facial Rejuvenation by Inducing Hyaluronic Acid Synthesis: A Pilot Preclinical Study.

Background: The level of dermal hyaluronic acid (HA) can be depleted by 75% at age 70. HA provides dermal hydration, volume, and thickness, making it a major component of the extracellular matrix. Restoration of dermal and epidermal HA can be achieved by combining radiofrequency (RF) energy and targeted ultrasound (TUS). The monopolar RF generates heat, with the TUS stimulating HA production. The heat induces a regenerative response in the skin, increasing the fibroblast activity and producing various extracellular matrix compounds, including HA. Objectives: To investigate the effect of the simultaneous application of RF + TUS or RF + US on the stimulation of HA production. Methods: Twelve animals underwent 4 treatments. Six were treated with transcutaneous RF + TUS and 6 with the combination RF + US. The opposite untreated side served as a control. Punch biopsies of the skin were taken at baseline, immediately posttreatment, 1 month, and 2 months posttreatment. The tissue was evaluated with real-time quantitative polymerase chain reaction (RT-qPCR), matrix-assisted laser desorption (MALDI) and time of flight (TOF), and confocal microscopy. Results: The RT-qPCR focused on assessing the production of has1 and has2, enzymes responsible for HA synthesis. RT-qPCR results of the RF + TUS group revealed a +98% and +45% increase in hyaluronic synthetase (HAS) 1 and HAS2 production after the treatments, respectively. The MALDI-TOF revealed a +224% increase in measured HA 2 months after the treatments. The changes were also visible in the confocal microscopy. The control group showed no significant (P > .05) results in either of the evaluation methods. Conclusions: Concurrent application of RF and TUS significantly enhances the natural regenerative processes in skin tissue.

First Evidence of Cutaneous Remodelling Induced by Synchronized Radiofrequency Aided by High-Intensity Facial Muscle Stimulation: Porcine Animal Model.

BACKGROUND: The quality of one's facial appearance diminishes with aging as skin and underlying soft tissues deteriorate. Connective tissue and musculofascial degeneration leads to skin laxity and wrinkles developing. OBJECTIVE: To evaluate the effects of synchronized radiofrequency with high intensity facial stimulation technology on dermal collagen and elastin fibers in a porcine model. MATERIALS AND METHODS: Eight sows were divided into Active (N = 6) and Control (N = 2) groups. Synchronized radiofrequency and high intensity facial stimulation were delivered to the ventrolateral abdomen. The Active group received four 20-minute treatments, once a week. Control group was untreated. Skin biopsy sample were histologically analyzed for connective tissue changes pre- and post-treatment. Data were analyzed statistically (α = 0.05). RESULTS: In the Active group: the collagen-occupied area at baseline was 1.12 ± 0.09 × 106 µm 2 and increased by +19.6% ( p < .001) at 1-month and by +26.3% ( p < .001) 2 months post-treatment; elastin-occupied area at baseline was 0.11 ± 0.03 × 106 µm 2 and increased by +75.9% ( p < .001) at 1-month and +110.8% ( p < .001) at 2-months follow-up. No significant changes ( p > .05) found in the Control samples. CONCLUSION: Collagen and elastin fiber content increased significantly after treatments. Connective tissue in the treatment area was denser up to 2-months post-treatment.

Novel Technology for Facial Muscle Stimulation Combined With Synchronized Radiofrequency Induces Structural Changes in Muscle Tissue: Porcine Histology Study.

BACKGROUND: With age, facial muscles lose the ability to complete contractions properly, resulting in limitation of facial expressions and fat shifting, and leading to skin creases and wrinkling. OBJECTIVES: The aim of this study was to determine the effects of the novel high intensity facial electromagnetic stimulation (HIFES) technology combined with synchronized radiofrequency on delicate facial muscles, using an animal porcine model. METHODS: Eight (n = 8, 60-80 kg) sows were divided into the active group (n = 6) and the control group (n = 2). The active group underwent four 20-minute treatments with radiofrequency (RF) and HIFES energies. The control group was not treated. Histology samples of muscle tissue were collected by a punch biopsy (6 mm in diameter) from the treatment area of each animal at baseline, 1-month, and 2-month follow-up. The evaluation included staining of the obtained tissue slices with hematoxylin and eosin and Masson's trichrome to determine the changes in muscle mass density, number of myonuclei, and muscle fibers. RESULTS: The active group showed muscle mass density increase (by 19.2%, P < .001), together with elevated numbers of myonuclei (by 21.2%, P < .05) and individual muscle fibers, which increased from 56.8 ± 7.1 to 68.0 ± 8.6 (P < .001). In the control group, no significant changes were seen in any of the studied parameters throughout the study (P > .05). Finally, no adverse events or side effects were observed in the treated animals. CONCLUSIONS: The results document favorable changes after the HIFES + RF procedure at the level of the muscle tissue, which may be of great importance in terms of maintenance of facial appearance in human patients.

Effects of oil-based adjuvants on the immune response of pigs after dermal administration of antigen and evaluation of the immunization level after a subsequent Actinobacillus pleuropneumoniae challenge in pigs.

Route of vaccine delivery can greatly impact the immunogenicity, efficacy and safety of the vaccine. Four groups of piglets were immunised transdermally (t.d.), intradermally (i.d.) or intramuscularly (i.m.) with the same doses of antigen in combination with a water-in-oil-in-water emulsion adjuvant Montanide™ ISA 201 VG or with a microemulsion adjuvant Montanide™ IMS 1313 VG N ST (Seppic, France). The last group was left without vaccination as a control group. All animals were subsequently exposed to the infection induced by Actinobacillus pleuropneumoniae (App). The immune response was evaluated with respect to the intensity of systemic and mucosal antibody formation, their isotype characterisation and rate of cell-mediated immunity. These findings were compared with the intensity of adverse local reactions and level of protection in experimental challenge. Monitoring of the local reaction at the injection site after each administration showed that microemulsion adjuvant IMS 1313 was less reactogenic than the water-in-oil-in-water emulsion ISA 201. In terms of efficacy, both dermal administrations were less immunogenic than the i.m route. The i.m. injection induced higher anti-App9 IgG and IgM titres. Nevertheless, IgG1 and IgG2 isotypes analysis revealed a close immunological profile between i.m. and i.d. routes. The concentration of IFN-γ from peripheral blood after in vitro restimulation with the specific antigen was only increased in the i.m. group at the day of challenge (D35) and two weeks after (D49). Interestingly, the smallest gross pulmonary lesions were observed in the i.d. vaccinated group (3.4%) compared to the control group (39.4%) and to groups with other routes of administration. Taken together, these results suggest that i.d. administration of vaccines is a promising approach. Even the i.d. vaccine was more reactogenic and slightly less immunogenic than the i.m. vaccine, its protection effectiveness seemed to be superior.

Vaccine against Streptococcus suis Infection in Pig Based on Alternative Carrier Protein Conjugate.

Streptococcus suis is a serious pathogen in the pig industry with zoonotic potential. With respect to the current effort to reduce antibiotic use in animals, a prophylactic measure is needed to control the disease burden. Unfortunately, immunization against streptococcal pathogens is challenging due to nature of the interaction between the pathogen and the host immune system, but vaccines based on conjugates of capsular polysaccharide (CPS) and carrier protein were proved to be efficient. The main obstacle of these vaccines is manufacturing cost, limiting their use in animals. In this work, we tested an experimental vaccine against Streptococcus suis serotype 2 based on capsular polysaccharide conjugated to chicken ovalbumin (OVA) and compared its immunogenicity and protectivity with a vaccine based on CRM197 conjugate. Ovalbumin was selected as a cheap alternative to recombinant carrier proteins widely used in vaccines for human use. We found that the ovalbumin-based experimental vaccine successfully induced immune response in pigs, and the IgG antibody response was even higher than after immunization with capsular polysaccharide-CRM197 conjugate. Protectivity of vaccination against infection was evaluated in the challenge experiment and was found promising for both conjugates.

Cytokine expression by CD163+ monocytes in healthy and Actinobacillus pleuropneumoniae-infected pigs.

Distinct monocyte subpopulations have been previously described in healthy pigs and pigs experimentally infected with Actinobacillus pleuropneumoniae (APP). The CD163+ subpopulation of bone marrow (BM), peripheral blood (PB) and lung monocytes was found to play an important role in the inflammatory process. The inflammation is accompanied by elevation of inflammatory cytokines. The aim of the study was to evaluate the contribution of CD163+ monocytes and macrophages to cytokine production during APP-induced lung inflammation. Cytokine production was assessed by flow cytometry (FC) and quantitative PCR (qPCR) in CD163+ monocytes and by qPCR, immunohistochemistry/fluorescence in lungs and tracheobronchial lymph nodes (TBLN). Despite the systemic inflammatory response after APP infection, BM and PB CD163+ monocytes did not express elevated levels of a wide range of cytokines compared to control pigs. In contrast, significant amounts of IL-1ß, IL-6, IL-8 and TNF-α were produced in lung lesions and IL-1ß in the TBLN. At the protein level, TNF-α was expressed by both CD163+ monocytes and macrophages in lung lesions, whereas IL-1ß, IL-6 and IL-8 expression was found only in CD163+ monocytes; no CD163+ macrophages were found to produce these cytokines. Furthermore, the quantification of CD163+ monocytes expressing the two cytokines IL-1ß and IL-8 that were most elevated was performed. In lung lesions, 36.5% IL-1ß positive CD163+ monocytes but only 18.3% IL-8 positive CD163+ monocytes were found. In conclusion, PB and BM CD163+ monocytes do not appear to contribute to the elevated cytokine levels in plasma. On the other hand, CD163+ monocytes contribute to inflammatory cytokine expression, especially IL-1ß at the site of inflammation during the inflammatory process.

MALDI MSI Reveals the Spatial Distribution of Protein Markers in Tracheobronchial Lymph Nodes and Lung of Pigs after Respiratory Infection.

Respiratory infections are a real threat for humans, and therefore the pig model is of interest for studies. As one of a case for studies, Actinobacillus pleuropneumoniae (APP) caused infections and still worries many pig breeders around the world. To better understand the influence of pathogenic effect of APP on a respiratory system-lungs and tracheobronchial lymph nodes (TBLN), we aimed to employ matrix-assisted laser desorption/ionization time-of-flight mass spectrometry imaging (MALDI-TOF MSI). In this study, six pigs were intranasally infected by APP and two were used as non-infected control, and 48 cryosections have been obtained. MALDI-TOF MSI and immunohistochemistry (IHC) were used to study spatial distribution of infectious markers, especially interleukins, in cryosections of porcine tissues of lungs (necrotic area, marginal zone) and tracheobronchial lymph nodes (TBLN) from pigs infected by APP. CD163, interleukin 1ß (IL­1ß) and a protegrin-4 precursor were successfully detected based on their tryptic fragments. CD163 and IL­1ß were confirmed also by IHC. The protegrin-4 precursor was identified by MALDI-TOF/TOF directly on the tissue cryosections. CD163, IL­1ß and protegrin­4 precursor were all significantly (p < 0.001) more expressed in necrotic areas of lungs infected by APP than in marginal zone, TBLN and in control lungs.