Functional hypothesis of the juxtaoral organ: Role of collagen types I and III.

The juxtaoral organ (JOO) was first described in human embryos by (Chievitz, 1885). In human adults, it has been described in the buccotemporal space—located within the soft tissue between the medial surface of the mandible and the buccinator muscle—innervated by the buccal nerve (Zenker, 1953). Histologically, the JOO consists of parenchyma formed by nests of non-keratinizing squamous epithelial cells surrounded by two layers or strata of connective tissue that form the stroma—the stratum fibrosum externum (SE) and the stratum fibrosum internum (SI). In addition, the JOO is rich in small nerve branches that form the stratum nervosum (Zenker & Salzer, 1961). Some reviews have reported epithelial islands residing in peripheral nerves similar to JOO, cases of hyperplasia, hamartomas or neuroectodermal tumours; even in biopsies, the JOO can be misinterpreted as carcinomas (Pantanowitz & Balogh, 2003; D’Andrea et al., 2015; Suárez-Quintanilla et al., 2020). The function of the JOO is not clearly defined. Despite numerous immunohistochemical studies on the JOO, there are no studies on the expression of collagen types I and III in its stroma (Table 1). Such information is crucial for its understanding since collagens play structural roles and contribute to mechanical properties, organization and shape of tissues (Ricard-Blum, 2011). Thus, the objective of this work was to carry out a histological study of JOO after dissection of the infratemporal region in adult human cadavers, analysing the immunoexpression of collagen types I and III and its disposition (by using the picrosirius red staining technique). For this study, sixteen formalin-fixed cadaver-head specimens were collected from the former Department of Anatomy and Embryology II and the Body Donation Centre of the Complutense University of Madrid. In all cases, a bilateral dissection of the infratemporal fossa was performed. In six cases, the JOO dissection was attempted, but it could not be identified macroscopically. In 10 cases, we explanted the region where the JOO was suspected to be. Therefore, the buccal nerve and surrounding tissue were resected, from its access to the pterygomandibular region until the buccinator muscle was reach (Figure 1). Histological examination allowed the JOO to be identified. The preparations were processed and routinely stained, including picrosirius red. Furthermore, immunohistochemistry on paraffin sections was performed following the protocol for the ready-to-use Vectastain (PK-7200). The following primary antibodies were used: mouse anti-collagen type I (sc-59772, 1:200) and rabbit polyclonal antibody anti-collagen type III (bs-0549R, 1:200). According to our findings, the JOO is a fusiform structure of variable length (6–11 mm) and thick (1–2 mm), surrounded by loose connective tissue (Figure 2a). The parenchyma of the JOO is formed by interconnected nests of epithelial cells, of different size and shape, with a multilobular appearance (Figure 2b-d). In some nests, the internal cells show a whorl-like or concentric arrangement and a larger cytoplasm, and other nests show small vesicles with amorphous material (Figure 2e). The stroma was formed by dense connective tissue that forms the capsule (SE), organized in concentric layers (Figure 2b,e,f), in which type I collagen predominated (Figure 2g). Inside the stroma, the SI was formed by loose connective tissue, with a predominance of type III collagen, arranged radially between the epithelial nests (Figure 2f,h). A range of immunohistochemical findings have been reported by immunohistochemical studies performed in different species and types of tissues, focused on the parenchyma or the nerves fibres (Table 1). However, there are no previous studies of the expression of collagen I and III in the stroma that emphasize its role as a mechanoreceptor. The role of the JOO stroma is important for its organization and morphological function. Previous studies suggested that the difference in the proportion of type I/III collagens, and the arrangement and size of the fibres are important characteristics for the mechanical resistance and elasticity of the tissues, since the elastic and dynamic properties are significantly different in both types of collagens (Asgari et al., 2017). We suggest that the SE formed predominantly by concentrically arranged collagen I, giving the JOO solidity and resistance to stretching and compression. The dense connective tissue that surrounds the small neurovascular bundles that reach the JOO serves to fix it to the neighbouring tissue. In addition, the SI principally formed by radially arranged collagen III provides the elastic properties to JOO. However, immunostaining of collagen type III is also slightly noticed in the SE. These results are consistent with the general distribution and function of collagen types (Gelse et al., 2003). Indeed, collagen type III is associated with collagen type I in many tissues (Kuivaniemi & Tromp, 2019). The importance of the JOO stroma during development has been reported previously. In rats, there is evidence that it is derived from the neural crest (Velasco et al., 2012). Furthermore, it has also been suggested that the melanin pigmentation identified within the JOO stroma in human adults is produced by melanocytes derived from the neural crest (Ide et al., 2003). Recently, it has also been reported that the stroma presents perineural characteristics (Suárez-Quintanilla et al., 2020). The JOO is located in the pterygomandibular region in relation to the buccal nerve. Its topographical relationships with the masticatory muscles, the buccinator muscle and the pterygomandibular raphe, as well as the organization of its stroma, suggest that JOO acts as a mechanoreceptor stimulated by actions such as chewing or swallowing. The knowledge of the topography and histology of the JOO can help to avoid misinterpretations related to the invasive front of carcinomas. Indeed, some microscopic features have been reported to be helpful in differentiating between the JOO and invasive perineural carcinoma (Müller, 2016; Pantanowitz & Balogh, 2003). In addition, this report also suggests that the immunoexpression of collagen types I and III in the stroma of the JOO seems to be consistent with its role as a mechanoreceptor. This work was partially supported by grant number PR87/19-22587 from Santander Bank/Complutense University of Madrid, to which we thank. However, the financial support has not influenced the results, and none of the authors discloses any conflict of interest. The authors also wish to sincerely thank those who donated their bodies to science so that anatomical research could be performed. Results from such research can potentially improve patient care and increase our overall knowledge. Therefore, these donors and their families deserve our highest gratitude. The authors declare no conflict of interest. Antonio José Mérida-García: Conceptualization; Formal analysis; Investigation; Writing-review & editing. Jorge Murillo-González: Funding acquisition; Investigation; Supervision; Writing-original draft; Writing-review & editing. Elena Martínez-Sanz: Formal analysis; Writing-original draft; Writing-review & editing. Javier Catón: Formal analysis; Investigation; Writing-original draft; Writing-review & editing. Luis A. Arráez-Aybar: Investigation; Writing-review & editing. José Martín-Cruces: Formal analysis; Writing-review & editing. Teresa Cobo: Formal analysis; Writing-review & editing. José Antonio Vega: Formal analysis; Supervision; Writing-review & editing. José Ramón Mérida-Velasco: Conceptualization; Formal analysis; Supervision; Writing-original draft; Writing-review & editing. All the procedures followed in this study were in accordance with the ethical standards of the Complutense University of Madrid (Spain) and in accord with the Helsinki Declaration. The peer review history for this article is available at https://publons.com/publon/10.1111/odi.13959. The authors declare that all the data of this study are available on request from the corresponding author, Jorge Murillo-González.