Preliminary Study of Yucatan Porcine Breast Morphology: Identifying Basic Differences and Similarities for Surgical Model Applications

Abstract

Background/Objectives: The porcine mammary anatomy is poorly characterized, and structures are difficult to differentiate macroscopically, unlike human mammary tissue. The objective of this exploratory study was to describe the Yucatan porcine breast tissue morphology and identify the basic differences and similarities to human breast tissue. Methods: Samples from an adult, non-parous female Yucatan pig were prepared utilizing various methods: freezing at −80 °F (−26.67 °C) with a thickness of 0.5 cm/post-fixation in formalin; freezing at −20 °F (−6.67 °C) with a thickness of 0.5 cm/post-fixation in formalin; or formalin fixed and incised at ~0.8 cm. A descriptive comparison of the gross and microscopic images of the porcine breast morphology to the previously described human breast anatomy was performed. Results: As examined grossly, frozen sections allowed narrower serial cross-sectioning and better visualization of the structures and relationships. The mammary glands were poorly demarcated with extensive interspersed adipose tissue throughout the periphery. The mammary tissue appeared grossly as pigmented tissue and extended within ~0.5 cm from the skin surface, ~2.0 cm deep (within ~0.5 cm of the deep muscle layer), and ~6.5 cm laterally (centered on a teat). There were a number of similarities between Yucatan porcine and human breast tissue, yet there were several inherent structural differences. In contrast to human mammary tissue, porcine mammary glands consist of more diffuse acinar tissue, less well demarcated by defined fascial, lamellar, and ligamentous structures. Conclusions: The Yucatan porcine mammary morphology and similarities to the human mammary region allow for the use of this animal model to guide those developing relevant technologies or performing local surgical interventions in the preclinical setting.

1. Introduction

Breast surgery is a commonly performed surgery in the United States, that is increasing in frequency. In 2023, there were approximately 304,000 breast augmentations and 157,000 breast reconstruction procedures performed, representing a 2% and 4% increase in the procedures since 2022 [1]. The rising number of breast surgeries can be attributed to the advancements in surgical techniques and the developments of new technologies, which create a demand for cosmetic and reconstructive procedures.

Devices have been and continue to be developed for breast surgery. Silicone implants were developed in the 1960s for use in esthetic and breast reconstruction procedures [2]. Scaffolds including acellular dermal matrices (ADMs) and synthetic mesh devices have been developed and used in breast surgery to support the implant and reinforce the soft tissue in the breast. The scaffolds provide critical structural support, especially in complex reconstructions and revision surgeries. The materials studied most frequently fall into three categories: biologic mesh (i.e., ADM; e.g., Alloderm^® [LifeCell Corporation, Branchburg, NJ, USA], Allomax^® [Bard Davol Inc., Warwick, RI, USA], Strattice^® [LifeCell Corporation, Branchburg, NJ, USA], SurgiMend^® [Integra LifeSciences Corporation, Plainsboro, NJ, USA], OviTex^® [Tela Bio Inc., Malvern, PA, USA]), absorbable synthetic mesh (e.g., Vicryl [Ethicon, Somerville, NJ, USA], TIGR^® [Novus Scientific, Uppsala, Sweden], Phasix™(Becton, Dickinson, and Company, Franklin Lakes, NJ, USA), GalaFLEX™ [Galatea Surgical, Lexington, MA, USA], DuraSorb^® [Surgical Innovation Associates, Chicago, IL, USA]), and nonabsorbable synthetic mesh (e.g., TiLoop^® [PFM Medical, Cologne, Germany], Seragyn™ [Serag-Wiessner KG, Naila, Germany]) [3]. The materials vary in their composition, resorption rates, and mechanical properties, allowing surgeons to choose the most appropriate scaffold.

With the evolution and increased utilization of devices in breast surgery, there is an increased need to have an animal model to assess the safety prior to use in humans. Animal models provide valuable insights into biological integration, mechanical performance, and the potential complications associated with new devices and technologies. Among the various animal models, porcine models are particularly advantageous given the similar animal and mammary size, physiology, disease progression, and absence of fur [4,5]. Porcine models are widely used in many areas including organ transplantation, emergency procedures, and tissue engineering [4]. The basic mammary gland anatomy and development in swine are extensively described in the veterinary literature [6,7]. Further, porcine models for free-flap breast reconstruction have been developed [8,9,10]. The anatomy of the latissimus dorsi muscle flaps (LDMFs), musculus rectus abdominis, and musculus transversus abdominis has been characterized, and the layout is similar to the location and anatomical features in humans making the pig a good model for breast reconstructive surgery [11,12]. The existing porcine models used for free-flap breast reconstruction were mostly utilized for training purposes and developing new surgical techniques [8,9,10]. The authors conclude that the porcine model is a good model and highlight the comparisons between swine and humans related to fat distribution, muscle differences, and differences in vascularity that required adjustments in surgical technique [8,9,10].

Despite the use of porcine models in breast reconstruction and for breast cancer [5], the porcine mammary anatomy is poorly characterized, and the structures are difficult to differentiate macroscopically. A Yucatan swine model is a clinically relevant model often chosen for its size and availability of adipose tissue. The Yucatan adipose more closely resembles human tissue compared to Yorkshire adipose tissue which is more dense and fibrotic [13]. While clinically relevant, the macroscopic anatomy differs greatly between the human and Yucatan swine models. The mammary and associated tissues are supported by the surgeon’s hand in Figure 1. The Yucatan mammary tissue shown in Figure 1 (left) does not have distinct anatomical features or differences in tissue morphology, as can be identified in a human’s breast tissue as shown in Figure 1 (right). A cadaveric study was performed to identify a suitable scaffold implantation site and outlined the layers of anatomical sites on a minipig grossly [14]. However, there is a need to understand the structures microscopically given that the anatomy is not easily visualized grossly.

Therefore, the objective of this exploratory study was to describe the Yucatan porcine breast tissue morphology and identify the basic differences and similarities to human breast tissue anatomy previously described.

2. Materials and Methods

Three mammary gland samples were collected from an ~55 kg cadaver adult (~15 months old), nulliparous, and non-parous female Yucatan pig that was sourced from a commercial laboratory animal vendor (Figure 2). The tissue samples were harvested immediately post-euthanasia, processed, and analyzed at CBSET, Inc. (Lexington, MD, USA) by a board-certified veterinary pathologist.

One sample was processed using each of the following methods to obtain serial transverse (i.e., medial-lateral) sections through the cranial–caudal length of the mammary gland; it was frozen (−80 °F [−26.67 °C]) then cut with a bandsaw at 0.5 cm, frozen (−20 °F [−6.67 °C]) then cut with a bandsaw at 0.5 cm, or formalin fixed and incised at ~0.8 cm. Macroscopic digital photos were taken of the serial sections from each sample. Frozen samples were then post-fixed in formalin.

The tissue regions of interest were sampled into oversized (e.g., 2 × 3”) cassettes, paraffin processed, microtome cut, and stained with hematoxylin and eosin (H&E) and Masson’s trichrome (MT) for histomorphologic evaluation. Sub-gross images were captured using a standard digital camera, and higher-magnification slide photomicrographs were captured using a standard microscope with a mounted digital camera.

3. Results

3.1. Gross Observations

As examined grossly, frozen sections were considered superior to fixed sections as they allowed both narrower serial cross-sectioning as well as better gross preservation, definition, and visualization of the anatomic structures and relationships. All the figures include images of samples frozen at −20 degrees Fahrenheit (−6.67 °C).

Regardless of preparation, the mammary gland appeared grossly as pigmented (gray) tissue arranged in a tubulovillous structure. The glands were poorly demarcated with extensive interspersed adipose tissue throughout the periphery. The mammary tissue extended within ~0.5 cm from the skin surface, ~2.0 cm deep (within ~0.5 cm of the deep muscle layer), and ~6.5 cm laterally (centered on a teat).

3.2. Histomorphology

As examined histologically, there were comparable results across the sample groups regardless of fixation/sectioning. All the samples exhibited excellent preservation of morphology and cellular features, with no distortion, disruption, artifact, or other issues. Representative samples are depicted in Figure 3 and Figure 4.

The mammary tissue is surrounded by the dermis on the ventral side and skeletal muscle and fascia on the dorsal side. The ventral side of the sample depicted by the dashed boxed area in Figure 3A,B shows the dermis and hypodermis present before the mammary tissue (Figure 3C). The mammary tissue is present between the dashed box and solid box depicted in Figure 3A,B. Figure 3B,C highlight radially oriented, thin linear collagen bundles comparable to Cooper’s ligaments in humans. Extending dorsally from the mammary tissue, there is skeletal muscle, adipose, fascia, adipose, and skeletal muscle identified (Figure 3D). The observations described were consistent for all the three samples from three different mammary glands, regardless of the processing methods.

Cellular features identified include the mammary duct represented by the solid black box in Figure 4B and depicted in Figure 4C. The approximate furthest extension of the mammary gland tissue was identified and is represented by the dotted blue box in Figure 4B and depicted in Figure 4D. The quiescent mammary gland acini separated by abundant adipose tissue is represented by the dashed red box in Figure 4B and depicted in Figure 4E. The cellular features described were consistent for all the three samples from three different mammary glands, regardless of the processing methods.

3.3. Porcine–Human Comparison

A descriptive comparison of the porcine breast morphology described in the current study and human breast anatomy previously described [9] is presented to show the basic similarities and differences between the structural and anatomical features for the porcine (Figure 5, left) and human (Figure 5, right) mammary anatomy. The fundamental morphology of the Yucatan swine is similar grossly and histologically. The mammary tissue is surrounded by fascia and lamellar fat anteriorly and posteriorly. The mammary tissue has lactiferous ducts and corpus mammae. Cooper’s ligament is present in humans and an equivalent was identified in the porcine mammary tissue. The lobule containing alveoli was present in the human mammary anatomy only. In contrast to the human mammary anatomy, the porcine mammary glands consisted of more diffuse acinar tissue, less well demarcated by defined fascial, lamellar, and ligamentous structures.

4. Discussion

The objective of this exploratory study was to describe the Yucatan porcine breast tissue morphology and identify the basic differences and similarities to the human breast tissue anatomy previously described. The utility of porcine models is well accepted across fields given the similarities in size, anatomy, and disease progression [4,5]. Veterinary textbooks describe the mammary gland anatomy and histology in swine [6,7]. Several studies have described porcine models for free-flap harvesting and transfer in breast reconstruction [6,7,8]. The anatomy of the latissimus dorsi muscle flaps (LDMFs), musculus rectus abdominis, and musculus transversus abdominis in pigs has been characterized [11] and have a similar location and anatomical features to those in humans [11,12]. Although porcine models were used in breast reconstruction procedures, the differences in the anatomy of mammary tissue is secondary to the other anatomical structures [6,7,8]. The layers of the mammary tissue have been described grossly [14]. However, there is a need to understand the porcine mammary tissue microanatomy to inform the developing interventional approaches and guide surgical procedures as there is increased utilization and development of devices in breast surgery.

Procedurally, it is difficult to differentiate and identify porcine mammary tissue structures and landmarks macroscopically, unlike the human breast anatomy (Figure 1). In the current study in Yucatan swine, the mammary tissue extended within ~0.5 cm from the skin surface, ~2.0 cm deep (within ~0.5 cm of the deep muscle layer), and ~6.5 cm laterally (centered on a teat). Importantly, the teat position can vary based on the breed and location (e.g., anterior, middle, posterior) [15]. The porcine mammary gland of Yucatan swine appeared grossly as pigmented (gray) tissue in the current study. The pigmentation differs based on the breed [16]. It is important to note that not all Yucatan swine have the gray pigmentation around/in their mammary tissue, as described here. In fact, there are many cases where the entire tissue plane has no pigmentation and is uniformly white, as shown in Figure 1, or areas with and without pigmentation within the same animal. Future studies should include more animals to better determine the strain characteristics and account for the individual differences.

While the morphology is difficult to differentiate grossly, the current study described the microanatomy and cellular features that were visualized following histological processing that can be helpful for surgical model applications. As examined histologically, there were comparable results regardless of the fixation/sectioning. All the porcine samples exhibited excellent histologic preservation of the microanatomy and cellular features, with no distortion, disruption, artifact, or other issues. The histologic and anatomic features described in this study were consistent for all the three samples from three different mammary glands, regardless of the processing methods. However, using frozen gross specimens allowed narrower serial cross-sectioning during trimming and, following standard formal fixation, paraffin embedding, and microtomy, a better comparison of the anatomic structures and relationships between the gross samples and slides.

While the porcine mammary anatomy was difficult to differentiate grossly, the current study advances the understanding of the porcine mammary morphology that has been characterized histologically [7,17] by providing a depiction and demonstration of additional key features including the lactiferous duct, corpus mammae, and Cooper’s ligament equivalent. The approximate furthest extension of mammary gland tissue is also shown for reference. Cheng (2021) previously summarized the layers in the mammary glands: skin, subcutaneous fat and glandular tissue, anterior rectus sheath, rectus abdominis muscle, posterior rectus sheath, and peritoneum [14]. The extended description and characterization of the porcine mammary morphology described in this study is critical given that it is impossible to articulate the levels and tissue planes grossly in the porcine mammary tissue.

A number of similarities were identified between the porcine breast morphology described in this study and the human breast anatomy previously characterized by Rehnke (2018) and others in the field [9]. The mammary tissue is surrounded by fascia and lamellar fat. In addition, the mammary tissue has lactiferous ducts and corpus mammae. The morphology was similar, but the structures were not as well defined in the porcine mammary glands. The basic comparison was limited by the involuted nature of the mammary gland in nulliparous swine.

The porcine breast morphology sufficiently corresponded to the human breast features, allowing for the development of future preclinical models of clinical relevance for surgical interventions. Previous studies highlight the utility of the porcine model for free-flap breast reconstruction [8,9,10], scaffold implantation in the breast [14], and fat grafting [13,14]. The key similarities to humans include the size, presence of key structural features, and fat distribution in the Yucatan species [9,10,13]. Some important differences for consideration when using a porcine model include the thinner subdermal fat [9,10] and thick dermis compared to those in humans [9]. The advantages of the porcine model include minimizing the number of animals needed given that a pig has eight plus teats allowing multiple treatment conditions per animal [14] and the ability to establish safety with a large animal model with a more similar anatomy than small animal models. The Yucatan strain, specifically, is clinically relevant given the adipose tissue [13].

One limitation of the current preliminary, exploratory study is the use of a single animal of the Yucatan strain. Therefore, this summary provides a starting point to inform preclinical surgical evaluations and determine whether further anatomical characterization is necessary. Further anatomical and histological characterization would require additional animals and a quantitative comparison using an appropriate statistical analysis. While this study demonstrates the basic model utility for the development of preclinical surgical models in breast surgery, a broad application of the results from the study is limited given the possibility of the relevant anatomical differences across the porcine strains. Individual anatomical variation and sporadic anomalies also must always be considered. Future studies should characterize the anatomy in additional animals to account for individual anatomical anomalies and other strains which may have different characteristics.

The morphology of the Yucatan porcine breast tissue in this paper contextualizes the knowledge of the porcine structural mammary anatomy grossly and at the microscopic level. The Yucatan porcine mammary structural morphology and similarities to the human mammary region allow for the use of this animal model to guide those developing the relevant technologies or performing local surgical interventions in the preclinical setting.