The efficacy of mesenchymal stromal cell-derived therapies for acute respiratory distress syndrome—a meta-analysis of preclinical trials

In critically ill patients, ARDS is a severe clinical syndrome with high morbidity and mortality [1]. The pathophysiological features of ARDS are characterized by diffuse alveolar damage, acute noncardiogenic lung oedema, and decreased functional lung volume [2]. Patients with moderate and severe ARDS are usually in need of intubated mechanical ventilation. If exacerbated, the patients need be put in a prone position; alternatively, if the patient is unresponsive to regular treatment, ECMO should be employed as a salvage therapy [3]. While mechanical ventilation can provide urgently needed respiratory support, it can cause volutrauma, atelectrauma, and biotrauma, all of which may accentuate the condition of patients with ARDS [4]. To date, there are no evidence-based pharmaceutical agents for ARDS or any treatments directly targeting the pathophysiology of ARDS [5]. Mesenchymal stromal cells (MSCs)—a type of pluripotent stem cell—were first found in the bone marrow. With antibacterial, immunomodulatory, and tissue and organ repair and regeneration characteristics, MSCs have been widely investigated as a potential therapy in different scenarios for ALI/ARDS in the last few decades [6]. MSCs may be effective for ALI/ARDS caused by a variety of pathogenic factors, as they can ameliorate lung permeability, decrease inflammatory cell infiltration, downregulate inflammatory mediators and upregulate anti-inflammatory mediators. The effects of MSCs were assumed to be due to their engraftment and proliferation, which were demonstrated to be rather limited [7]; however, according to current research, the paracrine- and endocrine-related secretomes are more important for tissue damage repair [8‐10]. MSCs may manage intracellular oxidative stress via exosomes, which can be engulfed and reutilized by macrophages, thus suppressing inflammation and regulating immunity; therefore, MSCs may have potential in lung injury treatment [11, 12]. However, MSCs may be oncogenic, triggering an immune response that, per se, may exacerbate ARDS in patients. Furthermore, the storage of MSCs may interfere with their gene expression or viability. As extracellular vesicles (EVs) are manufactured from conditioned medium (CM) by centrifugation, in this study, they are collectively referred to as MSC-derived therapies (MDTs). MDTs that contain these secretomes may have potential for treating ALI/ARDS. Today, MDT-related research is a fast-growing field [9, 13‐15]. Although MDTs cannot proliferate like MSCs, they have the advantages of easier preservation and transfer. Furthermore, in comparison with MSCs, MDTs have reduced immunogenicity and are thus an attractive solution for allogeneic transplants [16]. We will investigate the efficacy of MDT for ALI/ARDS to evaluate whether it can improve survival, lower lung injury severity, and regulate immune balance via a meta-analysis of animal models.

EVs comprising exosomes and microvesicles are extracted from conditioned medium (CM) through centrifugation; similar to CM, they also possess the MSC secretome [8, 36]. Thus, CM and EVs were integrated as MSC-derived therapies (MDTs) in this study, the purpose of which was to summarize the evidence of MDTs for ALI/ARDS. In our study, meta-analyses were conducted for parameters such as lung injury score (LIS), survival, neutrophils in BALF, protein in BALF, W/D ratio and inflammatory mediators. These thorough analyses were important and essential for demonstrating the efficacy of MDT for ALI/ARDS. To date, only a few similar studies regarding lung diseases have been conducted, and none of them have solely focused on ALI/ARDS [37, 38]. To our knowledge, this is the first meta-analysis focused on the efficacy of MDT for ALI/ARDS in preclinical studies.

Our meta-analysis demonstrated that MDT can mitigate the severity of ALI/ARDS in animal models. The lung injury score (LIS), a scoring scale under a microscope, is a widely used pathophysiological tool to assess lung injury severity in preclinical trials. In our study, the pooled result indicated that MDT significantly reduced LIS, which is direct evidence that MDT can attenuate lung injury severity. The results also suggested that, in animal trials, MDT was able to increase survival. Moreover, our study revealed that MDT can downregulate the levels of inflammatory factors such as IL-1, IL-6 and TFN-a while upregulating the level of IL-10, a well-known anti-inflammatory factor. Thus, MDT may regulate immunologic balance in a desirable manner. The immunomodulatory effects of MDT may be an important reason for ameliorated lung injury and improved survival.

The W/D ratio of the lung is an extensively utilized parameter to assess pulmonary vessel permeability in animal studies, which was demonstrated to be decreased in our study. This reduced ratio indicated that MDT can improve lung water clearance. Our meta-analysis suggests that MDT can downregulate the infiltration of neutrophils into the alveolar space. The decrease in neutrophils in alveoli not only attenuated inflammation and subsequent high vessel permeability in the lung but also reduced lung tissue damage, which in turn may improve the outcomes. In addition, our study discovered that the total protein in BALF was reduced with MDT treatment. The reduction in total protein was not just the consequence of downregulated lung vessel permeability but also may be the mechanism of improved lung compliance. As the “hyaline membrane” is a protein-rich fluid formed on the alveolar surface, it pathophysiologically increases alveolar interfacial tension and blocks oxygenation in ALI/ARDS.

EVs can be divided into exosomes, microvesicles, or apoptotic bodies depending on size, biogenesis, and composition. Exosomes are generally homogenous in size, with a diameter ranging from 40 to 200 nm, whereas microvesicles are relatively heterogeneous, ranging from 50 to 1000 nm in diameter up to the state of the cell during release [8, 39]. Furthermore, the process of vesicle formation and release from cells also differs between exosomes and microvesicles. Each of the EV subtypes has its own characteristic surface and intracellular markers. Although exosomes, microvesicles and conditioned medium may externally differ from one another, in our study, the available subgroup analyses of each subtype demonstrated that they were consistently efficacious for ALI/ARDS in preclinical trials. The probable reason for this consistency is that they all internally contain the therapeutic secretome which is irrelevant to size or formation. Specifically, exosome (EX), microvesicle (MV) and conditioned medium (CM) subgroup meta-analyses were available for outcomes such as LIS, neutrophil counting in BALF, total protein in BALF, IL-6 in the lung and IL-10 in the lung. Only the IL-6 subgroup meta-analysis detected statistically significant difference between the EX and CM subgroups. Whether this difference was generated by confounding factors or by original efficacious differences remains to be further studied. The detailed subgroup meta-analysis results can be found in the Additional file 1.

ARDS is a common clinical syndrome that causes respiratory distress due to refractory hypoxia for a variety of heterogeneous aetiologies. The hallmark of ARDS is noncardiogenic lung oedema, a result of diffuse alveolar damage, increased permeability of lung vessels, infiltration of inflammatory cells, and protein-rich fluid leakage into the alveolar space, which causes overwhelming hypoxia [2]. The popularity of lung protective ventilation [40, 41], mainly characterized by low tidal volume and low inspiratory pressure, decreased ARDS mortality in the early 2000s [42, 43]. Furthermore, in 2013, an RCT discovered that prone positioning can significantly reduce 28-d and 90-d mortality on the basis of lung protective ventilation [44]. The control of driving pressure was also associated with increased survival in ventilator settings in ARDS [45]. Other measures taken for respiratory support, such as lung recruitment and PEEP titration, may increase mortality; thus, they are not recommended in the clinical routine [46]. Although increased understanding of ARDS has been achieved in recent decades, no pharmaceutical agents have been verified as effective treatments. Trials for medications such as aspirin, intravenous salbutamol, recombinant human keratinocyte growth factor, rosuvastatin, and simvastatin were all ineffective because they did not result in reduced mortality of ARDS [5].

To date, with regard to treating ARDS, there is no targeted medicine that has proven to be effective [47]. Since 2007, a large body of preclinical trials have investigated the efficacy of MSC therapy for ALI/ARDS, demonstrating that MSCs can stabilize the alveolar-capillary barrier, enhance alveolar fluid clearance, and decrease infection and inflammation [48‐50]. Microvesicles derived from stem cells were reported to contain secretomes, such as protein and mRNA components that are crucial for stem cell renewal and expansion [51]. Since MSCs have been revealed to have the potential to treat ALI/ARDS, conditioned medium (CM) or extracellular vesicles (EVs) of MSCs, which possess these secretomes, have been the subjects of studies in recent years.

In basic numerical research, MSCs exhibit lung protective potential via paracrine growth and anti-inflammatory factors and downregulation of inflammatory pathways. Not only were MSC intensively investigated in vivo and in vitro in preclinical trials, several human trials, regarding the safety of MSC’s for ARDS, were also carried out in the past few years [52‐54]. Although the safety of MSCs has been questioned because of their oncogenic possibility, to date, no direct MSC-related adverse events have been detected in the above trials. The safety of MDT should be more reassuring since no live cells were transplanted during the treatment [55]. According to Katie Famous et al., clinically, ARDS can be divided into two subphenotypes, which have different inflammation statuses and respond differently to fluid infusion [56]. Whether these distinct ARDS subphenotypes respond differently to MSCs or MDTs is a topic worthy of future research.

There are several limitations in our meta-analysis. First, the overall sample size of our study was small due to the small sample size of preclinical trials. Second, the causes of ALI/ARDS were not unanimous within the studies. Third, the sources of MDT were not consistent within the studies; therefore, those of both human and animal origin were investigated. Additionally, the dosage of MDT and the intervention duration also varied among the studies. These limitations may generate substantial heterogeneity among the studies, which may thereby confound the results of our analyses. Finally, the lack of large animal trials and regular clinical parameters (such as respiratory mechanics) in the included MDT trials may miss some important information useful to guide its application in clinical settings.