Development of a Porcine Model of Delayed Wound Healing

• Chronic wounds do not proceed through the normal phases of wound healing and often stall in the inflammatory phase.
• Chronic wounds affect over 6 million people in the US due to slower wound healing associated with aged skin, impaired blood circulation, diabetes and other comorbidities [1]. 
• Rodent or porcine diabetic models are frequently used to model chronic wounds, such as diabetic ulcers, but these models are unable to fully replicate the clinical complications, especially chronicity of the wound. 
• In this study, we developed a model to mimic healing impairment that is caused by glycosylation of collagen resulting from extended exposure to hyperglycemia [2].

An In Vivo Efficacy Evaluation of Various Wound Management Products for Their Relative Debridement Activity, Using a Porcine Wound Eschar Model

Productive debridement is critical for the care and support of the healing of chronic wounds. However, there is a shortage of comparative data on the efficacy of various products and technologies with debridement potentials, currently available on the market. In order to expand the understanding of how different products can affect wound debridement, a study was conducted using a porcine, full-thickness burn model, which has previously been reported in the literature [Shi, L; et al 2009]

In this model, a series of twenty, 2 cm, full-thickness burn wounds were created on the backs of pigs. Immediately after the burns were created, a variety of different products were applied to the eschar using a split-back study design. Using a time0ciurse study design, debridement of the bum eschar was evaluated by clinical (visual) assessment. Depending on manufacturer’s recommendations, products were re-applied either every 24 hours or every 72 hours.

The results of this study demonstrate clear differences in the debridement activity of the various products tested, highlighting the utility of this animal model as a tool for screening debridement technologies in a pre-clinical setting.

Anatomical Effects in the Development of a Delayed Wound Healing Model

ACell, Inc. currently markets multiple configurations of its Urinary Bladder Matrix (UBM-ECM) product,  called MatriStem®. This material is derived from the decellularized basement membrane and tunica propria layers of the porcine bladder and consists of extracellular matrix proteins including collagens, glycosaminoglycans, and growth factors, which provide both a structural and biological framework for tissue healing [1, 2, 3].

Although the mechanism of action behind ECM-mediated constructive remodeling is not yet fully understood, it has been hypothesized that these ECM-derived biomaterials act as in situ bioactive regenerative templates, serving as substrates for progenitor cell infiltration and differentiation. The ECM biomaterial can be considered a concentrated version of the body's own natural scaffold that occurs after injury. The body is hypothesized to act as a bioreactor, providing additional site-specific biomechanical and biochemical cues that guide cell differentiation. Studies have demonstrated the chemo-attractive effects of UBM-ECM biomaterial breakdown products on progenitor cells   [4].

In June 2012, ACell, Inc. received a Space Medicine and Related Technologies Commercialization Assistance Program (SMARTCAP) award from the National Space Biomedical Research Institute (NSBRI) to develop a novel MatriStem UBM gel formulation, which can be easily administered to wounds in space. The development of a novel gel formulation of MatriStem UBM technology that preserves the bioactivity and vulnerary properties of the biomaterial would provide astronauts with a powerful new tool to combat lacerations and abrasions incurred during flight missions. Although existing MatriStem products have the potential to improve healing in space, current dressing configurations would be difficult to administer in low gravity and maintain position under high activity levels. Promising prototypes have been developed (Figure 1) and are undergoing continuing characterization. In order to mimic the delayed healing in space, a novel model of ischemic wound healing in rats has been developed which will be used to test the in vivo efficacy of the gel prototypes.

Staphylococcus strains show resistance to antibiotics in both in-vitro and in-vivo settings.

Hospitals often use in vitro methods to assess the susceptibility of bacterial strains and use the information collected to guide physicians in the selection of suitable antibiotics to treat infected patients. Much research has already shown that in vitro methods can lead to false conclusions of bacterial susceptibility since bacteria grown planktonically are hundreds or thousands of times more sensitive to antibiotic treatment compared to bacteria growing in a biofilm. For this study, we used a rat subcutaneous implant model to test this concept by directly challenging “sensitive” and “resistant” Staphylococcus strains first in vitro and then in vivo. As anticipated, both “sensitive” and “resistant” bacteria were immune to extremely high levels of antibiotic therapy when they have contaminated a device implant. These results confirm the necessity of using in vivo animal models such as this rat subcutaneous implant model for testing the efficacy of new antibiotics particularly when those antibiotics are being developed to treat device-related infections and/or antibiotic resistant organisms.

In Vivo-In Vitro Evaluation of Bacteria Aerosolization During Treatment with Acoustic Pressure Wound Therapy* for Infected Wounds

Acoustic pressure wound therapy (APWT)* delivers acoustic pressure waves to the wound bed via a gentle, sterile saline mist to remove slough, fibrin, tissue exudates, and bacteria.1 This study was designed to determine whether treatment with APWT2 results in hazardous levels of bacteria aerosolization into the treatment environment during treatment of infected wounds and what effect, if any, universal precautions would have on reducing or eliminating aerosolized microbial exposure.

Development Of a Regenerative Wound Dressing For Improved Healing In Space

ACell, Inc. currently markets multiple configurations of its Urinary Bladder Matrix (UBM-ECM) product, called MatriStem®. This material is derived from the decellularized basement membrane and tunica propria layers of the porcine bladder and consists of extracellular matrix proteins including collagens, glycosaminoglycans, and growth factors, which provide both a structural and biological framework for tissue healing [1,2].

Although the mechanism of action behind ECM-mediated constructive remodeling is not yet fully understood, it has been hypothesized that these ECM-derived biomaterials act as in situ bioactive regenerative templates, serving as substrates for progenitor cell infiltration and differentiation. The ECM biomaterial can      be considered a concentrated version of the body's own natural scaffold that occurs after injury. The body is hypothesized to act as a bioreactor, providing additional site-specific biomechanical and biochemical cues    that guide cell differentiation. Several studies have demonstrated the chemo-attractive effects of UBM-ECM biomaterial breakdown products on progenitor cells.

Evaluation of CHG Compatibility of skin Care Products in an Ex Vivo Porcine Dermal Model

The use of preventative measure to reduce healthcare-associated infections, including the use of the antimicrobial chlorhexidene gluconate (CHG)1, is becoming more widespread. It is known that under certain circumstance, commonly used components of skin care products can reduce the antimicrobial effectiveness of CHG2-5. Although review of ingredients has been recommended, this does not provide definite guidance for the clinician. Therefore, an ex vivo porcine skin model was used at an independent microbiology laboratory to test the CHG compatibility of three skin care products using methods that simulated clinical usage while allowing assessment of CHG antimicrobial activity.

Complex biofilms show variability of bacteria survival over long durations: implications for in vitro screening of antimicrobial actives.

Development of new antimicrobials has not kept pace with the rise in drug resistant bacterial infections seen today, requiring clinicians to use multiple antibiotics in larger doses, for longer periods of time. Modified Robbins Devices (MRDs), designed with small uniform sampling coupons have greatly increased the efficiency in testing novel agents against biofilms, known to play a critical role in drug resistance and chronic infections. Due to the slow metabolism of bacteria in biofilms, long exposure times are often required to demonstrate efficacy of antimicrobial agents. However, in order to establish the relevancy of long exposure times, it is important to first determine how long control (untreated) biofilms can survive without the benefit of nutrient media, which can interfere with the function of some antimicrobial agents. This is further complicated when the biofilm is composed of a mixed culture biofilm with more than on microorganism. In this study, we grew a complex biofilm matrix composed of S. aureus, P. aeruginosa, and E. coli. After the biofilm reached maturity, it was removed from nutrient media and submerged in a non-nutritive buffered saline solution (PBS) for up to 36 hours. Total and species-specific colony forming unit (CFU) counts were determined using both selective and non-selective agar plates. The results of this study have important implications for the testing of antimicrobial and anti-biofilm agents.

Crosslinking Agents for the Development of an Infected Chronic Wound Model

The development and characterization of chronic wound models in animals have historically been an active area of research. Based on the established correlation of both wound duration and biofilm with wound chronicity, we have set out to challenge and expand the crosslinked chronic wound model by adding bacterial infection (S. epidermidis, P. aeruginosa, and Fusobacterium in a 1:1:0.5 ratio). 

We had hypothesized that the cytotoxicity of the crosslinkers would present a barrier to bacterial survival. But contrary to our hypothesis, persistence of the species was observed and the data showed that the glutaraldehyde and EDC delayed healing better than genipin or ribose and maintained high bioburden of clinically relevant bacteria.

Retrospective Analysis of nearly 300 Wounds from Porcine Wound Healing Studies Performed over 6 Years

Pigs are one of the most heavily utilized models for the development and testing of novel drugs and dressings in the field of wound healing. As a contract laboratory specializing in wound care, BRIDGE PTS, Inc. has a large body of historical data on wound healing rates in Yorkshire pigs with in a specific range of age and size. The purpose of this analysis was to compare the data from nearly 300 control wounds from multiple wound healing studies performed over a roughly 6-year period. Data from these studies was collated and overall trends were delineated. The analysis shows that,in this model, a clearly recognizable healing curve can be observed: with the inflammatory phase occurring from Days 0 to 4, the regenerative phase occurring from Days 4 to 14, and the remodeling phase occurring from Day 14 onward. Extrapolated rates of change for these different periods are also reported. This data could be useful in future studies for the verification of normative wound behavior and as a baseline for comparison with novel wound healing agents or dressings designed to accelerate or decelerate wound healing.

Biofilm Longevity Assessment of E. coli, S. aureus, and C. albicans in Nonnutritive Phosphate Buffered Saline

The Modified Robbins Device (MRD) remains an important tool in the study of biofilms and is routinely used to assess the antimicrobial properties of novel agents on biofilms. As such, the natural lifespan of biofilms (e.g. nutrient depletion, osmotic pressure, etc.) must be controlled for when testing for biofilm reduction caused by exposure to anti-biofilm agents using this testing apparatus. We assessed the longevity of three mono-culture biofilms of different host organisms (Escherichia coli (ATCC 25922), Staphylococcus aureus (ATCC 6538), and Candida albicans (ATCC 10231)) in nonnutritive phosphate buffered saline (PBS) incubated in 6-port static testing devices.  The biofilms were established in a MRD on sterile PVC plastic sampling coupons.  After 22 hours of incubation at 37°C ± 3°C in half-strength Tryptic Soy Broth (TSB; bacterial test systems) or half-strength Yeast Dextrose Broth (YD/B; C. albicans) circulating at 60 mL/hour, via a peristaltic pump, the sampling coupons were transferred from the MRD to a 6-port static sampling device filled with PBS.  Coupons were then sampled to identify the concentration of viable microbes remaining on the coupon.  At 7 days (168 hours), the survival rates of these microorganisms varied widely and not in an easily predictable fashion. These results show the importance of testing biofilm survival independent of test articles since there is a large range in biofilm longevity for different microbial species.