Characteristics of laryngotracheal stenosis

Laryngotracheal stenosis (LTS) is a broad term encompassing narrowing of the airway at the level of the larynx, subglottis, or trachea. Laryngotracheal stenosis (LTS) is a rare but severe condition. Besides the functional impairments as a result of airway obstruction, stenosis of the airways leads to considerable morbidity and mortality. [1,2] The main cause of LTS is intubation. [3] Current options for treating stenosis involve endoscopic dilation, laser surgery, laryngotracheal reconstruction, or life-long tracheostomy, but are often suboptimal as new scar tissue frequently develops leading to restenosis. [3,4] It has been suggested that LTS develops because of altered fibroblast responsiveness to anti-fibrotic signals during mucosal repair leading to excessive production of fibrosis. [5] Furthermore, there seems to be an altered inflammatory response leading to hypertrophic scars and a correlation between early inflammatory reaction to injury of the mucosa and the degree of scarring. [6]

The initial, preferred and less invasive method of treatment is endoscopic dilation, but this method has only proven success rates in paediatric patients of 64%; not incorporating the need for repetitive treatments due to re-stenosis and accompanying morbidity. [3,4] Alternatively, laryngotracheal reconstruction is required for severe stenosis. [3,7] This procedure holds high success rate but entails severe risks and subsequent morbidity. [6]

Mesenchymal stem cells

Mesenchymal stem cells (MSCs) are adult multipotent stem cells characterized by their adherence to plastic, their surface antigen expression, and their ability to differentiate into various connective tissue lineages including adipogenic, chondrogenic, myogenic and osteogenic cells. [8,9]

MSCs have during the last decades received extensive interest due to their potential therapeutic use as treatment for multiple diseases. [10] MSCs have shown anti-inflammatory and immunosuppressive properties, an ability to migrate to the exact site of injury, and a capacity to secret soluble factors crucial for cell survival and proliferation with minimal side effects. [10,11] Furthermore, MSCs currently indicate promising results in regenerative medicine. [10] In addition, MSCs are easily accessible for isolation most commonly from the bone marrow (BM-MSC) or adipose tissue (ASC) and have shown great expansion potential supporting the prospective of MSCs as a therapeutic agent. It has been shown that ASC increase wound healing with less scarring in skin ulcers [12] and the prospect of using ASC for therapeutic purpose in treating LTS is promising.

This study systematically evaluated the literature on the effects of MSCs on the conductive airway. We aimed to clarify the potential of MSCs in the conducting airways in regard to potentially treating stenosis and to evaluate their potential in a future human trial.

Systematic literature search and eligibility criteria

In January 2017, one author (KKJ) systematically searched the PubMed, EMBASE, Google Scholar, Cochrane Library, and clinicaltrials.org for articles in the English and Scandinavian languages. We included studies evaluating the effect of exogenous supplied MSC on the conductive airway in animals regardless of the publication date. Only studies concerning the conductive airways (trachea to the bronchioles) were included. Studies concerning the effector-mechanisms by which MSCs exert their properties were excluded as were studies evaluating the effect of MSCs on the airway in combination with other therapeutic regimes. Finally, studies concerning the effect of MSCs on the alveolar function, lung parenchyma and vascular bed were excluded. Due to conflict of interests the study done by Macchiarini, P. et al. on clinical transplantation of a tissue-engineered airway was excluded.

The following keywords (MeSH terms included in PubMed) were used: mesenchymal stromal cell, or MSC, or mesenchymal stem cell, adipose derived stromal cells, or adsc preadipocytes, or processed lipoaspirate cells, or stromal vascular fraction cells and Airways when searching PubMed and Embase. In Google Scholar, clinicaltrials.org and Cochrane Library the search were performed with the keywords “stem cell” and “airways”.

The following data were extracted from the included studies: study design, study participants, graft donor, source of graft, intervention, control groups, origin of stem cell, statistical tests.

The effect of MSCs on inflammation in the airway

Five studies (n = 153) evaluated the effect of MSCs on inflammation in the airways, (Table 1). [14–18] All studies used a preclinical model of induced asthma with either ovalbumin or toluene diisocyanate (TDI) as an inflammatory sensitizer (Table 1). [14–18]

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Table 1. Overview of studies: MSCs and inflammation in the airways.

https://doi.org/10.1371/journal.pone.0185283.t001

Three of the studies (n = 100) administered MSCs after sensitization [15,17] while two studies (n = 53) [14,18] administered the MSCs before the sensitization. [18] All studies used BM-MSCs and administrated the MSCs via intravenous injection.

All studies could demonstrate a reduction in inflammation in the MSC treated group compared to the control group (Table 1). This was either measured as a significant decrease in the amount of inflammatory cells peribronchially [14,15,18] a reduction in the total white blood cell count, or a significant reduction in neutrophiles and eosinophils in blood samples [16,18]. Finally two studies demonstrated a significant reduction in the level of pro-inflammatory cytokines in the circulation in the MSC treated group compared to the control group. [14,17] In three of the studies the MSCs group was also demonstrated to have a significant reduction in goblet cell hyperplasia compared to the control group. [15,16,18]

Furthermore, two studies demonstrated a significant reduction in the thickness of the epithelium, the subepithelial smooth muscle layers, and the basement membrane in the upper airways. [15,17]

The effect of MSCs on fibrosis in the airways

Two of the above studies (n = 32) also addressed MSCs effect on fibrosis and collagen deposit in the upper airway (Table 1). [16,18]

One study on induced asthma was not able to demonstrate a significant change in fibrosis after administration of MSCs, evaluated by the amount of collagen deposition. [16] Another study on induced asthma [18] demonstrated an increase in collagen deposition following the sensitizer. However, administering MSCs caused that the amount of collagen deposition, only reached the level in the sham group.

The use of MSCs in tissue repair

Six studies (n = 103) investigated the use of MSCs as a means to induce tissue repair, (Table 2). [7,19–23]

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Table 2. Study results of the effect of MSCs on tissue engineering.

https://doi.org/10.1371/journal.pone.0185283.t002

One study (n = 30) [20], investigated the participation of MSCs in the recovery of injured airway epithelium. Naphthalene was used to induce lung damage one month after MSC transplantation into the jugular vein. It was observed that BM-MSCs could adhere to the airways and form patches of epithelial lining in the conductive airways. [20]

Two studies (n = unknown) evaluated the repopulation of decellularized lung scaffold with MSCs for subsequent clinical transplantation. [21,22] Mendez JJ. et al. [21] showed attachment to the airways when using MSCs derived from adipose tissue. In regard to epithelization, Mendez JJ. et al. showed that MSCs from both bone marrow and adipose tissue underwent epithelial cell differentiation.

This contrasts to Daly AM. et al. [22] that repopulated their scaffolds with BM-MSCs but their cells did not differentiate into pseudostratified epithelia.

Three studies (n = 73) evaluated the repair of a tracheal defect by surgically replacing the damaged area with a tracheal graft or a scaffold. [7,19,23] Ott LM. et al. [7] and Gray FL. et al. [19] used patch-type decellularized scaffold to repair a tracheal defect and investigated if seeding of MSCs onto the scaffold would be beneficial for the tracheal repair, measured by survival rate, severity of the stenosis, the epithelialization, and histology of the scaffold. Go, T. et al. (n = 30) [23] investigated the engineering of a functional long-segment graft and replaced six cm of the trachea with a tracheal graft. The tracheal defects were made on the lamb fetus, and after the defect was repaired the fetus was returned to the uterine cavity.

Ott LM. et al. found that the survival rate was higher in the BM-MSC group compared to the scaffold-only group and had a higher level of immature cartilage which could indicate that BM-MSCs contribute to cartilage formation.

In regards to cross section area and preventing stenosis no clear results exist. Ott LM. et al. observed a larger cross section area in the scaffold-only group compared with the scaffold seeded with BM-MSC. Gray FL. et al. did not show a significant difference in cross section area between the two groups. Gray FL. et al. argued that the scaffolds were made from tracheal segments of rabbits and were thus not originally programmed to grow or dilate. Go T. et al. showed that MSC-derived chondrocytes seeded on the external surface resulted in less stenosis of the tracheal graft. Go T. et al. [23] continued showing the beneficial effect of epithelialization which help prevent bacterial/fungal contamination.