Search Results (24)

Anastomotic leaks after colorectal resection are serious surgical complications. We have compared the integrity of two common colorectal anastomosis techniques, end-to-side (ES) and end-to-end (EE), to control specimens using a novel experimental setup that mimics anastomotic air leak tests, which are typically performed during surgeries. Freshly harvested porcine colonic sections from 23 F1 cross-species pigs were used. Pressure measurements and video imaging were used to monitor the ex vivo experiments on EE, ES, and Control specimens. Using EE (n = 16), ES (n = 12), and Control (n = 22) specimens, leak pressure was 282.6 ± 3.0 mm Hg for EE, 282.8 ± 2.6 mm Hg for ES, and 294.4 ± 12.1 for the Control. Time to leakage was 106.3 ± 28.1 s for EE, 263.9 ± 2127.0 s for ES, and 194.5 ± 90.2 s for the Control. We found that, while EE and ES have nearly identical leak pressures, ES was superior in terms of time to leakage and tissue expansion, which may explain why ES anastomoses have a lower clinical anastomotic leak rate. Two dependent variables representing stress and strain of colonic tissues were introduced. These variables showed ES was comparable to the Control. The experiments were simulated successfully using the finite element method (FEM). This research provides a reproducible ex vivo system with a corresponding FEM system to study the differences between anastomosis techniques and may help design anastomoses with lower leak rates and improve patient outcomes in colorectal surgeries. Full article

Many colonic surgeries include colorectal anastomoses whose leaks may be life-threatening, affecting thousands of patients annually. Various studies propose that mechanical interaction between the staples and neighboring tissues may play an important role in anastomotic leakage. Therefore, understanding the mechanical behavior of colorectal tissue is essential to characterizing the reasons for this type of failure. So far, experimental data characterizing the mechanical properties of colorectal tissue have been few and inconsistent, which has significantly limited understanding their behavior. This research proposes an approach to developing an anisotropic hyperelastic material model for colorectal tissues based on uniaxial testing of freshly harvested porcine specimens, which were collected from several age- and weight-matched pigs. The specimens were extracted from the same colon tract of each pig along their circumferential and longitudinal orientations. We propose a constitutive model combining Yeoh isotropic hyperelastic material with fibers oriented in two directions to account for the hyperelastic and anisotropic nature of colorectal tissues. Experimental data were used to accurately determine the model’s coefficients (circumferential, R² = 0.9968; longitudinal, R² = 0.9675). The results show that the proposed model can be incorporated into a finite element model that can simulate procedures such as colorectal anastomoses reliably. Full article

Adeno-associated virus (AAV) vectors are among the most widely used delivery vehicles for in vivo gene therapy as they mediate robust and sustained transgene expression with limited toxicity. However, a significant impediment to the broad clinical success of AAV-based therapies is the widespread presence of pre-existing humoral immunity to AAVs in the human population. This immunity arises from the circulation of non-pathogenic endemic human AAV serotypes. One possible solution is to use non-human AAV capsids to pseudotype transgene-containing AAV vector genomes of interest. Due to the low probability of human exposure to animal AAVs, pre-existing immunity to animal-derived AAV capsids should be low. Here, we characterize two novel AAV capsid sequences: one derived from porcine colon tissue and the other from a caprine adenovirus stock. Both AAV capsids proved to be effective transducers of HeLa and HEK293T cells in vitro. In vivo, both capsids were able to transduce the murine nose, lung, and liver after either intranasal or intraperitoneal administration. In addition, we demonstrate that the porcine AAV capsid likely arose from multiple recombination events involving human- and animal-derived AAV sequences. We hypothesize that recurrent recombination events with similar and distantly related AAV sequences represent an effective mechanism for enhancing the fitness of wildtype AAV populations. Full article

One hundred Actinobacillus pleuropneumoniae (App) and sixty Pasteurella multocida subsp. multocida serogroup A (PmA) isolates were recovered from porcine pneumonic lungs collected from eight central or southern states of Brazil between 2014 and 2018 (App) or between 2017 and 2021 (PmA). A. pleuropneumoniae clinical isolates were typed by multiplex PCR and the most prevalent serovars were 8, 7 and 5 (43, 25% and 18%, respectively). In addition, three virulence genes were assessed in P. multocida isolates, all being positive to capA (PmA) and kmt1 genes, all negative to capD and toxA, and most of them (85%) negative to pfhA gene. The susceptibility of both pathogens to tildipirosin was investigated using a broth microdilution assay. The percentage of isolates susceptible to tildipirosin was 95% for App and 73.3% for PmA. The MIC₅₀ values were 0.25 and 1 μg/mL and the MIC₉₀ values were 4 and >64 μg/mL for App and PmA, respectively. Finally, a multiple-dose protocol of tildipirosin was tested in suckling piglets on a farm endemic for both pathogens. Tildipirosin was able to prevent the natural colonization of the tonsils by App and PmA and significantly (p < 0.0001) reduced the burden of Glaesserella parasuis in this tissue. In summary, our results demonstrate that: (i) tildipirosin can be included in the list of antibiotics to control outbreaks of lung disease caused by App regardless of the capsular type, and (ii) in the case of clinical strains of App and PmA that are sensitive to tildipirosin based on susceptibility testing, the use of this antibiotic in eradication programs for A. pleuropneumoniae and P. multocida can be strongly recommended. Full article

Surgical site infections (SSIs) are among the most frequent healthcare-associated infections, resulting in high morbidity, mortality, and cost. While correct hygiene measures and prophylactic antibiotics are effective in preventing SSIs, even in modern healthcare settings where recommended guidelines are strictly followed, SSIs persist as a considerable problem that has proven hard to solve. Surgical procedures involving the implantation of foreign bodies are particularly problematic due to the ability of microorganisms to adhere to and colonize the implanted material and form resilient biofilms. In these cases, SSIs may develop even months after implantation and can be difficult to treat once established. Locally applied antibiotics or specifically engineered implant materials with built-in antibiotic-release properties may prevent these complications and, ultimately, require fewer antibiotics compared to those that are systemically administered. In this study, we demonstrated an antimicrobial material concept with intended use in artificial vascular grafts. The material is a silicone–hydrogel interpenetrating polymer network developed earlier for drug-release catheters. In this study, we designed the material for permanent implantation and tested the drug-loading and drug-release properties of the material to prevent the growth of a typical causative pathogen of SSIs, Staphylococcus aureus. The novelty of this study is demonstrated through the antimicrobial properties of the material in vitro after loading it with an advantageous combination, minocycline and rifampicin, which subsequently showed superiority over the state-of-the-art (Propaten) artificial graft material in a large-animal study, using a novel porcine tissue-implantation model. Full article

Background: Machine perfusion (MP) is increasingly used for human transplant organ preservation. The use of MP for research purposes is another opportunity for this technology. The porcine pancreas and liver are similar in anatomical size and function to their human counterparts, making them an excellent resource for research, but they have some important differences from human organs which can influence their research use. In this paper, we describe a technique developed and tested for the retrieval of porcine organs for use in research on perfused viable organs. Methods: Whole-organ porcine pancreata and livers were harvested at a commercial abattoir, following standard slaughterhouse processes. The standard slaughterhouse process involved a thoracotomy and mid-line laparotomy, and all the thoracoabdominal organs were removed. The pancreas, fixed in the retroperitoneum, was carefully dissected from its attachments to the surrounding structures, and tissue planes between the pancreas, spleen, duodenum, and colon were meticulously identified and dissected. Vessel exposure and division: The aorta, portal vein (PV), hepatic vein (HV), and hepatic artery (HA) were dissected and isolated, preserving the input and output channels for the liver and pancreas. A distal 3 cm of the aorta was preserved and divided and served as the input for the pancreas perfusions. The liver, PV, HV, and HA were preserved and divided to preserve the physiological channels of the input (PV and HA) and output (HV) for the liver perfusions. The porcine hepatic and pancreas anatomy shares significant resemblance with the gross anatomy found in humans, and this was taken into consideration when designing the perfusion circuitry. The porcine pancreas and spleen shared a common blood supply, with branches arising from the splenic artery. The organs were flushed with cold, heparinised normal saline and transported in a temperature-regulated receptacle maintained at a core temperature between 4 and 8 °C, in line with the standards of static cold storage (SCS), to a dedicated perfusion lab and perfused using our novel perfusion machine with autologous, heparinised porcine blood, also collected at the abattoir. Full article

Glaesserella parasuis (Gps), Gram-negative bacteria, are a universal respiratory-disease-causing pathogen in swine that colonize the upper respiratory tract. Highly Pathogenic Porcine Reproductive and Respiratory Syndrome Virus (HP-PRRSV2HP-PRRSV2) and Gps coinfections are epidemics in China, but little is known about the influence of concurrent coinfection on disease severity and inflammatory responses. Herein, we studied the effects of secondary HP-PRRS infection on clinical symptoms, pathological changes, pathogen load, and inflammatory response of Gps coinfection in the upper respiratory tract of piglets. All coinfected piglets (HP-PRRSV2 + Gps) displayed fever and severe lesions in the lungs, while fever was present in only a few animals with a single infection (HP-PRRSV2 or Gps). Additionally, HP-PRRSV2 and Gps loading in nasal swabs and blood and lung tissue samples was significantly increased in the coinfected group. Necropsy data showed that coinfected piglets suffered from severe lung damage and had significantly higher antibody titers of HP-PRRSV2 or Gps than single-infected piglets. Moreover, the serum and lung concentrations of inflammatory cytokines (TNF-α, IL-1β, IL-6, and IL-8) were also significantly higher in coinfected piglets than in those infected with HP-PRRSV2 or Gps alone. In conclusion, our results show that HP-PRRSV2 promotes the shedding and replication of Gps, and their coinfection in the upper respiratory tract aggravates the clinical symptoms and inflammatory responses, causing lung damage. Therefore, in the unavoidable situation of Gps infection in piglets, necessary measures must be made to prevent and control secondary infection with HP-PRRSV2, which can save huge economic losses to the pork industry. Full article

Annulus fibrosus (AF) damage is proven to prompt intervertebral disc (IVD) degeneration, and unrepaired AF lesions after surgical discectomy may boost herniation of the nucleus pulposus (NP) which may lead to further compression of neural structures. Moreover, vascular and neural ingrowth may occur within the defect which is known as a possible reason for discogenic pain. Due to a limited healing capacity, an effective strategy to repair and close the AF defect is necessary. In this study, using electrospinning technology, two nature polymers, silk fibroin and gelatin, were linked to imitate the unique lamellae structure of native AF. Our findings revealed that a multilayer electrospun-aligned fibroin/gelatin scaffold with mechanical and morphological properties mimicking those of native AF lamellae have been developed. The average diameter of the nanofiber is 162.9 ± 38.8 nm. The young’s modulus is around 6.70 MPa with an ultimate tensile strength of around 1.81 MP along preferred orientation. The in vitro test confirmed its biocompatibility and ability to maintain cell viability and colonization. Using a porcine model, we demonstrated that the multilayer-aligned scaffold offered a crucial microenvironment to induce collagen fibrous tissue production within native AF defect. In the implant-repaired AF, H&E staining showed homogeneous fibroblast-like cell infiltration at the repaired defect with very little vascular ingrowth, which was confirmed by magnetic resonance imaging findings. Picrosirius red staining and immunohistochemical staining against type I collagen revealed positively stained fibrous tissue in an aligned pattern within the implant-integrated site. Relative to the intact control group, the disc height index of the serial X-ray decreased significantly in both the injury control and implant group at 4 weeks and 8 weeks (p < 0.05) which indicated this scaffold may not reverse the degenerative process. However, the results of the discography showed that the effectiveness of annulus repair of the implant group is much superior to that of the untreated group. The scaffold, composed with nature fibroin/gelatin polymers, could potentially enhance AF healing that could prevent IVD recurrent herniation, as well as neural and neovascular ingrowth after discectomy surgeries. Full article

Regeneration of articular cartilage remains challenging. The aim of this study was to increase the stability of pure bioactive glass (BG) scaffolds by means of solvent phase polymer infiltration and to maintain cell adherence on the glass struts. Therefore, BG scaffolds either pure or enhanced with three different amounts of poly(D-L-lactide-co-glycolide) (PLGA) were characterized in detail. Scaffolds were seeded with primary porcine articular chondrocytes (pACs) and human mesenchymal stem cells (hMSCs) in a dynamic long-term culture (35 days). Light microscopy evaluations showed that PLGA was detectable in every region of the scaffold. Porosity was greater than 70%. The biomechanical stability was increased by polymer infiltration. PLGA infiltration did not result in a decrease in viability of both cell types, but increased DNA and sulfated glycosaminoglycan (sGAG) contents of hMSCs-colonized scaffolds. Successful chondrogenesis of hMSC-colonized scaffolds was demonstrated by immunocytochemical staining of collagen type II, cartilage proteoglycans and the transcription factor SOX9. PLGA-infiltrated scaffolds showed a higher relative expression of cartilage related genes not only of pAC-, but also of hMSC-colonized scaffolds in comparison to the pure BG. Based on the novel data, our recommendation is BG scaffolds with single infiltrated PLGA for cartilage tissue engineering. Full article

The objective of this paper is to present our findings on the biomechanical aspects of the virgin passive anisotropic hyperelasticity of the porcine colon based on equibiaxial tensile experiments. Firstly, the characterization of the intestine tissues is discussed for a nearly incompressible hyperelastic fiber-reinforced Holzapfel–Gasser–Ogden constitutive model in virgin passive loading conditions. The stability of the evaluated material parameters is checked for the polyconvexity of the adopted strain energy function using positive eigenvalue constraints of the Hessian matrix with MATLAB. The constitutive material description of the intestine with two collagen fibers in the submucosal and muscular layer each has been implemented in the FORTRAN platform of the commercial finite element software LS-DYNA, and two equibiaxial tensile simulations are presented to validate the results with the optical strain images obtained from the experiments. Furthermore, this paper also reviews the existing models of the active smooth muscle cells, but these models have not been computationally studied here. The review part shows that the constitutive models originally developed for the active contraction of skeletal muscle based on Hill’s three-element model, Murphy’s four-state cross-bridge chemical kinetic model and Huxley’s sliding-filament hypothesis, which are mainly used for arteries, are appropriate for numerical contraction numerical analysis of the large intestine. Full article

Clinically used heart valve prostheses, despite their progress, are still associated with limitations. Biodegradable poly-ε-caprolactone (PCL) nanofiber scaffolds, as a matrix, were seeded with human endothelial colony-forming cells (ECFCs) and human induced-pluripotent stem cells-derived MSCs (iMSCs) for the generation of tissue-engineered heart valves. Cell adhesion, proliferation, and distribution, as well as the effects of coating PCL nanofibers, were analyzed by fluorescence microscopy and SEM. Mechanical properties of seeded PCL scaffolds were investigated under uniaxial loading. iPSCs were used to differentiate into iMSCs via mesoderm. The obtained iMSCs exhibited a comparable phenotype and surface marker expression to adult human MSCs and were capable of multilineage differentiation. EFCFs and MSCs showed good adhesion and distribution on PCL fibers, forming a closed cell cover. Coating of the fibers resulted in an increased cell number only at an early time point; from day 7 of colonization, there was no difference between cell numbers on coated and uncoated PCL fibers. The mechanical properties of PCL scaffolds under uniaxial loading were compared with native porcine pulmonary valve leaflets. The Young’s modulus and mean elongation at F_max of unseeded PCL scaffolds were comparable to those of native leaflets (p = ns.). Colonization of PCL scaffolds with human ECFCs or iMSCs did not alter these properties (p = ns.). However, the native heart valves exhibited a maximum tensile stress at a force of 1.2 ± 0.5 N, whereas it was lower in the unseeded PCL scaffolds (0.6 ± 0.0 N, p < 0.05). A closed cell layer on PCL tissues did not change the values of F_max (ECFCs: 0.6 ± 0.1 N; iMSCs: 0.7 ± 0.1 N). Here, a successful two-phase protocol, based on the timed use of differentiation factors for efficient differentiation of human iPSCs into iMSCs, was developed. Furthermore, we demonstrated the successful colonization of a biodegradable PCL nanofiber matrix with human ECFCs and iMSCs suitable for the generation of tissue-engineered heart valves. A closed cell cover was already evident after 14 days for ECFCs and 21 days for MSCs. The PCL tissue did not show major mechanical differences compared to native heart valves, which was not altered by short-term surface colonization with human cells in the absence of an extracellular matrix. Full article

Tryptophan (Trp) plays an important role in piglet growth. However, the effect of dietary Trp on microbial flora is still poorly understood. A total of 40 28-d weaned piglets were allocated to four groups with 10 barrows per group and one pig per replicate. Piglets were fed a corn and soybean meal-based diet with 0.14%, 0.21%, 0.28%, or 0.35% Trp for four weeks. Five piglets from each diet group were euthanized, and blood and tissue samples were collected. The average daily body weight gain, average daily feed intake, feed conversion ratio, spleen index, pancreas index, longissimus dorsi muscle index, plasma insulin, 5-hydroxytryptamine, kynurenine, and Trp concentrations of weaned piglets increased in a dose-dependent manner (p < 0.05). Compared with the 0.14% Trp diet, the adequate-Trp diets (0.21%, 0.28%, or 0.35%) down-regulated the relative abundances of 12 genera including Turicibacter, Prevotella, Mitsuokella, Anaerovibrio, Megasphaera, Succinivibrio, Sutterella, Desulfovibrio, and Methanobrevibacter (p < 0.05); up-regulated the abundances of Ruminococcaceae, Lactobacillus, and Muribaculaceae in the colon (p < 0.05); and augmented the mRNA level and concentration of porcine β-defensin 2 in the small intestinal mucosa (p < 0.05). Moreover, Trp-adequate diets increased the abundances of Trp hydroxylase, indoleamine 2,3-dioxygenase, porcine β-defensin 2, phosphorylated mammalian target of rapamycin, and phosphorylated protein kinase B in the small intestinal mucosa (p < 0.05). We noted that a corn and soybean meal-based diet with 0.35% Trp may be a nutritional strategy to improve growth performance, intestinal mucosal barrier integrity, and intestinal microbial ecology in weaned piglets. Full article

The large intestinal epithelium is confronted with the necessity to adapt quickly to varying levels of oxygenation. In contrast to other tissues, it meets this requirement successfully and remains unharmed during (limited) hypoxic periods. The large intestine is also the site of bacterial fermentation producing short-chain fatty acids (SCFA). Amongst these SCFA, butyrate has been reported to ameliorate many pathological conditions. Thus, we hypothesized that butyrate protects the colonocytes from hypoxic damage. We used isolated porcine colon epithelium mounted in Ussing chambers, incubated it with or without butyrate and simulated hypoxia by changing the gassing regime to test this hypothesis. We found an increase in transepithelial conductance and a decrease in short-circuit current across the epithelia when simulating hypoxia for more than 30 min. Incubation with 50 mM butyrate significantly ameliorated these changes to the epithelial integrity. In order to characterize the protective mechanism, we compared the effects of butyrate to those of iso-butyrate and propionate. These two SCFAs exerted similar effects to butyrate. Therefore, we propose that the protective effect of butyrate on colon epithelium under hypoxia is not (only) based on its nutritive function, but rather on the intracellular signaling effects of SCFA. Full article

Knowledge of pathogenic potential, frequency and antimicrobial resistance patterns of porcine Streptococcus (S.) spp. other than S. suis is scarce. Between 2016 and 2020, altogether 553 S. spp. isolates were recovered from clinical specimens taken from Austrian swine stocks and submitted for routine microbiological examination. Antimicrobial susceptibility testing towards eight antimicrobial substances was performed using disk diffusion test. All isolates from skin lesions belonged to the species S. dysgalactiae subspecies equisimilis (SDSE). S. hyovaginalis was mainly isolated from the upper respiratory tract (15/19) and S. thoraltensis from the genitourinary tract (11/15). The majority of S. suis isolates were resistant to tetracycline (66%), clindamycin (62%) and erythromycin (58%). S. suis isolates from the joints had the highest resistance rates. S. suis and SDSE isolates resistant to tetracycline were more likely to be resistant to erythromycin and clindamycin (p < 0.01). Results show that different species of Streptococcus tend to occur in specific body sites. Nevertheless, a statement whether these species are colonizers or potential pathogens cannot be given so far. High resistance rates of S. suis towards tetracyclines and erythromycin and high recovery rates of S. suis from lung tissue should be considered when treating pigs with respiratory diseases. Full article

A microrobot system comprising an untethered tumbling magnetic microrobot, a two-degree-of-freedom rotating permanent magnet, and an ultrasound imaging system has been developed for in vitro and in vivo biomedical applications. The microrobot tumbles end-over-end in a net forward motion due to applied magnetic torque from the rotating magnet. By turning the rotational axis of the magnet, two-dimensional directional control is possible and the microrobot was steered along various trajectories, including a circular path and P-shaped path. The microrobot is capable of moving over the unstructured terrain within a murine colon in in vitro, in situ, and in vivo conditions, as well as a porcine colon in ex vivo conditions. High-frequency ultrasound imaging allows for real-time determination of the microrobot’s position while it is optically occluded by animal tissue. When coated with a fluorescein payload, the microrobot was shown to release the majority of the payload over a 1-h time period in phosphate-buffered saline. Cytotoxicity tests demonstrated that the microrobot’s constituent materials, SU-8 and polydimethylsiloxane (PDMS), did not show a statistically significant difference in toxicity to murine fibroblasts from the negative control, even when the materials were doped with magnetic neodymium microparticles. The microrobot system’s capabilities make it promising for targeted drug delivery and other in vivo biomedical applications. Full article