Burn injuries are a major health concern worldwide and a primary cause of illness and death on a global level. Standard treatment for burns typically involves the use of dressings and topical agents that establish a safeguard and facilitate the healing of wounds. Yet, the use of MSCs has emerged as a viable and optimistic substitute for treating burns. This study showed that MSCs have the potency to be used therapeutically to cure rat skin burns. Furthermore, results have shown that local transplantation of MSCs into burn wounds can significantly improve wound healing and skin regeneration, which could have important implications for treating burn injuries in humans.

Initially, the culture of BM-MSCs consisted of a diverse group of cells, while subsequent passages of the culture predominantly exhibited fibroblastic cells. These findings are consistent with previous research conducted on bovine (Corradetti et al. 2013), caprine (Pratheesh et al. 2014), canine (Csaki et al. 2009), and humans (Klemmt et al. 2011).

Nevertheless, our findings regarding the characterization of stem cells in cattle differ from the results reported in other studies (Rossi et al. 2014), where round and fibroblastic cells persisted throughout the culture, even at different stages. This study utilized CD34, CD73+, CD90+, and CD105+ surface antigen markers to assess all three BM-MSCs, consistent with the guidelines established by the ISCT, despite the use of additional marker subsets in other studies (Dominici et al. 2006).

For the best results in wound healing, it is necessary to have a coordinated and effective sequence of three phases that overlap with each other: inflammation, proliferation, and remodeling (Gurtner et al. 2008). Growth factors, cytokines, and chemokines mediate a wide range of cellular and molecular activities. These findings reveal that MSCs significantly enhanced wound healing rates. On the 14th day after the incision, there was a significant improvement (p < 0.001) in wound contraction in the groups that received MSC treatment compared to the control group. Similarly, wound contraction at days 21 and 28 was obviously (p < 0.001) better in the MSC-treated groups. Also, histopathological examination showed better results for the healed wound in MSC groups (in terms of collagen deposition, neovascularization, epithelization, and collagen assembly) than in the control group. MSCs have been proposed to enhance tissue repair through both paracrine signaling and their ability to differentiate.

While MSC paracrine signaling pathways control local cellular responses to injury, MSC differentiation contributes through the regeneration of damaged tissue (Hocking and Gibran 2010). Endogenous MSCs, such as those seen in skin sheaths and hair follicle bulges, are essential for the healing of wounds (Liu et al. 2014). These cells are distributed across the skin’s many niches, which are primarily classified into epidermal and dermal niches. MSCs play a crucial role in creating a conducive microenvironment for coordinated cellular and molecular activities, such as cell migration. This is achieved through the secretion of factors, macrovesicles, and exosomes by the MSCs (Hocking and Gibran 2010).

Recent research suggests that during the cultivation phase, MSCs release various substances essential for the cell’s regular physiological activity. Additionally, through a procedure called licensing, the innate MSCs are attracted to the wound site and stimulated by the inflammatory environment as they engage with the immune system (Shi et al. 2012) and begin to produce growth factors and cytokines resulting in compositional changes of local cytokines that are conducive to wound healing and tissue regeneration processes (Tamama and Kerpedjieva 2012). In our recent study, we observed a rapid increase in the levels of pro-inflammatory cytokines (TNF and IL-6) in response to short-term inflammation. However, we also found that BM-MSCs significantly counteracted these changes by quickly increasing in number.

Through paracrine mechanisms, wound healing improves by inhibiting the increasing angiogenesis, inflammatory process, collagen production, and stimulating fibroblast migration (Tamama and Kerpedjieva 2012). Furthermore, their paracrine factors suppress nucleic acid, protein metabolism, and apoptotic genes while increasing homeostatic and anti-apoptotic genes (Wu et al. 2006).

After severe burn damage, the systemic inflammatory response triggers the hypermetabolic response, which turns on protein breakdown and catabolism (Jeschke et al. 2008). As a result, pro-inflammatory mediators are released out of control, which worsens organ dysfunction and protein loss (Jeschke et al. 2002; Yeh et al. 2002). The failure of the organ’s performance increases the chances of contracting an infection and sepsis, ultimately failing multiple organs and eventual demise (Agarwal et al. 2023). This vicious circle is not well comprehended and challenging to overcome. There is a lack of testing for treatments that can alter the body’s response to burns, such as inflammation, hypermetabolism, or organ damage, in humans. While we have established a cytokine expression pattern in children who have sustained burns, further research is needed to find effective solutions (Finnerty et al. 2006); in animal research models used for burn pathophysiology, little information is known about the cytokine cascade’s duration or amplitude following an intense thermal injury.

After skin injury, a series of collaborative and dynamic processes occur in a well-coordinated manner to repair and regenerate the skin’s protective ability. These processes involve hemostasis, inflammation, proliferation, and remodeling (Reinke and Sorg 2012). After treatment with BM-MSCs, indicators of wound healing, such as the acceleration of re-epithelialization and thickness of the regenerated epidermis, were noted (Fu et al. 2006). To promote regeneration following burn injury, BM-MSCs are implanted into the damaged area and interact with the epithelial cells while transdifferentiating (Seppanen et al. 2013).

The investigation revealed that BM-MSCs can expedite the healing of skin wounds by promoting the production of collagen bundles, fibroblasts, basal cell proliferation, and vascularization, as well as exhibiting anti-inflammatory properties.

The results obtained by stereological analysis show that using BM-MSCs promotes the formation of the fibrous, thick dermis and granular tissue during epithelialization, increases collagen synthesis, and reduces the inflammatory cell numbers in the wound area.

Moreover, the investigation demonstrated that significant enhancement in vessel length and wound closure area was observed on days 21 and 28 following the introduction of burn injury, as compared to the group treated with BM-MSCs and other groups. The utilization of MSCs boosts the number of basal cells, fibroblasts, blood vessels, and fibrous tissue while reducing the count of inflammatory cells (neutrophils and lymphocytes), thereby facilitating more effective wound healing. Our histological observations are in close alignment with those of the study by Aryan et al. (2018). These findings provide strong evidence that MSCs can promote the proliferation of fibroblasts, which leads to intensified granulation tissue formation, the accumulation of collagen fibers, increased blood vessel formation, and improved re-epithelialization, contributing to enhanced wound healing.

The healing process of wounds consists of three phases—inflammation, proliferation, and remodeling—which occur concurrently and overlap with each other (Cañedo-Dorantes and Cañedo-Ayala 2019).

The extent and comprehension of the inflammatory reaction to burns in mice are limited. Moreover, the expression of cytokines in the blood plasma or serum of uninfected burned rodents is not well established. The typical cytokines measured after a burn injury in mice are IL-1, IL-6, or TNF, and the available literature presents conflicting results regarding the expression of these substances. In one particular study, mice with burns were found to have elevated serum levels of IL-6 and IL-1b after 24 h (Ipaktchi et al. 2006). On the basis of a subsequent inquiry, TNF, IL-6, and IL-10 were not detected in mice that had been burned or those that had undergone a sham procedure, during the first and seventh days following the burn (Murphy et al. 2005).

In contrast, we observed the presence of the three cytokines during days 7 and 14 after the burn was initiated. The number of burns in these two investigations differs significantly from ours; both experiments were conducted on mice that burned more than 25% (Murphy et al. 2005), 30% (Ipaktchi et al. 2006), and 35% total body surface area (TBSA) in this case. Patients’ inflammatory responses vary with burn size (Jeschke et al. 2007). Therefore, these variations in burn size could cause an apparent discrepancy in the data. The patterns we show for rats are supported by other research on rats that burnt more than 20% TBSA. Less than 72 h after burns, elevated levels of IL-1, TNF, and IL-6 have been noted (Kataranovski et al. 1999); our results found the same elevations in rats with burns. Rats burnt with above 60% TBSA showed an increase in IL-1b, IL-6, and IL-10 within hours of burn damage but no change in TNF, according to Gauglitz et al. (2008). Nonetheless, several cytokines that are not commonly evaluated in mice with burns have not been previously documented in terms of their adjustment after burns (Gauglitz et al. 2008).

The present study aimed to achieve two objectives. Firstly, to evaluate the cytokine expression in the serum of rats that were burned and compare it to the cytokine expression in normal, non-burned control rats and rats that underwent local transplantation of MSCs. Secondly, to assess the cytokine profile in burned rats that experienced altered expression after the burn injury, as these cytokines play a crucial role in the proliferation, differentiation, and clonal expansion of immune cells. Furthermore, these cytokines attract immune cells to the injury site. However, the massive upregulation of both pro-inflammatory and anti-inflammatory cytokines may cause non-specific inflammation, rather than a well-planned systemic inflammatory response where these two types of cytokines regulate each other in a coordinated manner (Li et al. 2020).

MSCs could modulate the immune system and reduce inflammation by inhibiting the production of pro-inflammatory cytokines such as TNF and IFN while increasing the secretion of anti-inflammatory cytokines such as IL-10 and IL-4. This property, combined with their inhibitory effect on neutrophil infiltration and IL-6, facilitates successful wound healing through the resolution of inflammation. In studies involving burnt animals, the injection of BM-MSCs significantly reduced the levels of IL-6 and TNF. The current study on a deep second-degree burn also reported an increase in TGFβ expression (Aggarwal and Pittenger 2005; Caliari-Oliveira et al. 2016; Gilbert et al. 2016).

In addition to their inhibitory effect on neutrophil infiltration and IL-6, the immunomodulating property of MSCs enables them to directly reduce the inflammatory response by inhibiting the production of pro-inflammatory cytokines like TNF and IFN and increasing the secretion of anti-inflammatory cytokines like IL-10 and IL-4. Successful wound healing is achieved through the resolution of inflammation. As a result, after subcutaneous injection of BM-MSCs in the burnt animals, the rate of production IL-6 and TNF was considerably reduced. We found an upregulation of TGF-β expression, which was reported in the current study of a deep second-degree burn.

Owing to their capacity to activate dermal fibroblasts, which increase the production of collagen type I and alter gene expression (Smith et al. 2010), MSCs speed up the healing of wounds (Rodrigues et al. 2019). In the current study, it was found that MSCs can hinder scarring by releasing VEGF and HGF, as well as by controlling the equilibrium between TGF1 and TGF3 through their paracrine signaling, as reported by (Colwell et al. 2005). Hsp90 plays a crucial part in this process by boosting cell survival and motility, and growth factors are not the only variables involved in wound closure (Li et al. 2012; Dong et al. 2016). Furthermore, Cheng et al. (2008) and Abdallah et al. (2023) found that it was accountable for the relocation of human epidermal and dermal fibroblasts. As for the process of epithelization, BM-MSCs enhanced this mechanism by promoting the proliferation of resident epidermal cells with the aid of EGF, or through their capacity to transform into epidermal cells (Kataoka et al. 2003; Rodrigues et al. 2019).

Our research reveals the concurrent involvement of multiple cytokines in the rat’s response after burns, marking the first instance of such a phenomenon. Additionally, we provide evidence indicating that six out of 11 cytokines exhibit comparable expression patterns throughout the inflammatory response to burns in rats, also a novel finding. Differences in the post-burn inflammatory response between mice, humans, and rats could be attributed to variations in species as well as the extent and severity of the burn. Moreover, we demonstrate that the duration and magnitude of the inflammatory response to burns in rats are like those documented in humans by Cui et al. (2023).

Previous research has shown that local injection of MSCs along with growth factors can affect the levels of pro-inflammatory and anti-inflammatory cytokines in the serum of rats that have undergone burn injury and local transplantation (Liu et al. 2020; Abdel-Gawad et al. 2021). The present study, on the other hand, focused on the impact of developmental factors injected in MSCs, which led to a significant decrease in the levels of pro-inflammatory cytokines such as TNFα and IL-6, while elevating the levels of anti-inflammatory cytokines, specifically IL-10 and TGF-β, in the serum of rats with burn injuries.

In prior research, skin injuries that received human umbilical cord MSCs transplantation exhibited a notable decrease in the number of inflammatory cells and pro-inflammatory cytokines when compared to the group that received PBS treatment (Liu et al. 2014).

Moreover, the rise in cytokines that have an anti-inflammatory effect, such as IL-10 and TGF-β, might be attributed to the transformation of MSCs into cell types, like fibroblasts, which are crucial in the restoration and regeneration of tissues. Furthermore, the secretion of growth factors, such as VEGF, that stimulate the development of new blood vessels and tissue regeneration, could occur because of the differentiation of MSCs into fibroblasts.

According to our research, one of the significant outcomes was the capacity of MSCs to stimulate the generation of growth factors, such as VEGF and TGFβ, which play a crucial role in tissue repair and wound healing. Another important finding was that the increased production of these growth factors, as observed in the histological analysis of treated wounds, accelerated the healing process and improved tissue regeneration (Oryan et al. 2019).

Additionally, we observed a reduction in inflammation and modulation of the immune response following local injection of MSCs, which is critical in the pathogenesis of burn injuries. This outcome implies that MSCs could potentially have a significant immunomodulatory effect, which could contribute to the improved healing observed in our experiments.

The potential of MSCs for medicinal and regenerative purposes has made them highly sought-after. MSCs can differentiate and repair damaged epithelium through differentiation and fusion, as well as release a diverse range of growth factors and cytokines (Li et al. 2023). Utilizing MSCs-conditioned medium (CM), which contains a wealth of growth factors and cytokines, is advantageous as it eliminates the risk of transplant rejection (Csaki et al. 2009). However, there is currently limited information regarding the comparison of MSC-CM in wound healing (Saadh et al. 2023).

In brief, the findings of this study indicate that providing growth factor-infused MSCs directly to burned rats’ injury sites can shift the balance of pro-inflammatory and anti-inflammatory cytokines in their blood, resulting in improved tissue healing and reduced inflammation. However, further investigation is required to determine the optimal dosage and timing of MSC injection to maximize therapeutic benefits.

To summarize, the study suggests that injecting MSCs with growth factors locally can be a useful therapy for burn wounds, as it reduces inflammation and promotes tissue repair. The decrease in pro-inflammatory cytokines such as TNF-α and IL-6 may be due to the immunomodulatory properties of MSCs, which suppress the activation and proliferation of immune cells like T, B, and natural killer cells. Moreover, MSCs release anti-inflammatory cytokines such as IL-10 and TGF-β, which suppress the production of pro-inflammatory cytokines.