DISCUSSION
We have verified that the AR mouse model was successfully established.
We first showed that intranasal delivery of MSCs-Exo could not only
relieve the main AR symptoms, sneezing and runny nose, but also decrease
various inflammation indicators, including the spleen index, tissue
staining, and the expression of inflammation-related cytokines.
Moreover, we found that the optimal concentration of MSCs-Exo was
4×108/mL. Besides, we demonstrated that the MAPK and
NF-κB pathways play an important role in inflammation, and that MSCs-Exo
ameliorated AR by inhibiting the expression of MAPK pathway proteins,
which modulated the immune response in the AR mouse model.
MSCs exert immunoregulatory effects by inhibiting T cell proliferation,
inactivating allogeneic T cells, influencing T cell
apoptosis22, and inhibiting B cell proliferation to
decrease antibody secretion23. Furthermore, MSCs
inhibit the differentiation and maturation of dendritic cells, resulting
in a decline in the ability to activate T cells24.
MSCs exert the above immunomodulatory effects mainly through the
paracrine secretion of exosomes25. The presence of
microvilli and columnar cells in the nasal cavity can intensify drug
absorption26, and intranasal administration is not
only safe but also convenient to operate. These advantages increase its
feasibility for clinical application. Therefore, we chose to study the
therapeutic effect of MSCs-Exo on AR by intranasal delivery; our results
verify that intranasal administration of
MSCs-Exo works not only locally
but also systemically.
After exposure to the specific
allergen, IgE mediates mast cell degranulation, causing itching and
sneezing and further promoting hemangiectasis and increasing secretion
of submucosal glands, thus resulting in obstruction and rhinorrhea. The
immunohistological characteristics of AR include the infiltration of
eosinophils and the predominant expression of cytokines secreted by Th2
cells, with a reduced activity level of Th1 cells, which contributes to
the late-phase response such as nasal obstruction and
hyperreactivity27, 28. Our results correspond with the
mechanism underlying AR described above. In the AR model, IgE, OVA-IgE,
IgG1, and histamine levels in BALF and serum and Th2 cytokine levels
increased, corresponding to severe local and systematic symptoms. After
intranasal administration of MSCs-Exo, the decrease in IgE, OVA-IgE, and
histamine levels relieved the classic symptoms; the decrease in CCL-11
levels is linked to lower eosinophil infiltration; the change in ICAM-1
and Th2 cytokine levels is linked to reduced inflammation; and increased
IFN-γ levels can stimulate the expression of IgG2a and inhibit the
production of IgG3, IgG1, IgG2b, and IgE29, thus
further exerting a therapeutic effect on AR. Many studies have also
demonstrated that after intravenous treatment with different kinds of
exosomes in an asthma mouse model, inflammation of lung tissues
decreased, and Th2 cytokine and inflammatory cytokine levels were lower
in serum and BALF30-32. All these results imply that
intranasal delivery of MSCs-Exo has an effect similar to that of
intravenous delivery to suppress allergic reactions.
In order to pave the way for clinical application in the near future, we
examined the effects of different doses, and found that the optimal
concentration is 4×108/mL. When the concentration of
MSCs-Exo is lower than 4×108/mL, the efficacy
decreases significantly. Once the concentration is higher than
4×108/mL, the curative effects do not further increase
significantly. The data show that MSCs-Exo works in a dose-dependent
manner, which supports future clinical research.
The activation of NF-κB plays an important role in allergic diseases,
inducing the accumulation of inflammatory cells33.
When activated by inflammatory signals, the released NF-κB induces the
transcription of many inflammatory cytokines, which are related to the
pathogenesis of asthma34. MAPKs are involved in many
different cellular events, including allergic
diseases35. Mammalian MAPKs are mainly divided into
three categories: extracellular signal-regulated kinase (ERK), c-Jun
N-terminal kinase (JNK), and p3836. The activation of
MAPK is critical for the production of inflammatory
cytokines37, and functional differentiation into Th1
or Th2 subsets38. Notably, inhibition of ERK promotes
the transition of Th2 lymphocytes to Th139. Our
results showed that the expression of genes related to the NF-κB and
MAPK pathways increased significantly in AR mice, but after MSCs-Exo
treatment, the expression of genes related to the NF-κB pathway did not
decrease significantly, while MSCs-Exo treatment significantly inhibits
ERK, JNK, JUN, and FOS expression. The results confirm that the NF-κB
and MAPK pathways play important roles in AR and indicate that MSCs-Exo
do not reduce the activation of NF-κB to alleviate the symptoms of AR.
MSCs-Exo inhibit inflammation by restricting OVA from affecting the MAPK
pathway in AR mice.
While our results have demonstrated that intranasal delivery of MSCs-Exo
is an effective therapy for AR, the specific mechanism has not been
adequately revealed. It has not been researched how the MAPK pathway
regulates the expression of cytokines to play a therapeutic role in AR.
The specific functional components of MSCs-Exo have not been identified,
and it is unknown how long they remain active. Previously, the effects
of intranasal and intravenous delivery of MSCs-Exo on the brain were
compared, and intranasal delivery was more
effective40. For AR, no direct comparisons between
intravenous and intranasal delivery have been made. Note also that we
only studied intranasal administration of MSCs-Exo isolated from human
umbilical cord; and while this extends the current treatment methods, it
will be interesting to test exosomes secreted by other cells to examine
the different effects.
Intranasal administration of MSCs-Exo has been researched in various
diseases, including complete spinal cord injury41,
microglia-mediated neuroinflammation42, and autism
spectrum disorders43. It is noteworthy that intranasal
delivery of MSC-Exo could substantially expand pulmonary IL-10-producing
interstitial macrophages to protect against allergic asthma in
mice44. However, research on intranasal delivery of
MSC-Exo in AR is still scarce. Our results offer a theoretical and
experimental basis for the future clinical local application of MSCs-Exo
in the nasal cavity for the treatment of AR, which can effectively
alleviate pain in AR patients and has significant clinical and social
value.