Dexmedetomidine Reduces IL-4 and IgE Expression Through Downregulation of TLR4/NF-κB Signaling Pathway to Alleviate Airway Hyperresponsiveness in OVA Mice

Background: Asthma is a disease that affects health worldwide. It is characterised by inammation and airway hyperreactivity. Because airway hyperreactivity can occur in other diseases, perioperative airway hyperreactivity is more insidious and widespread than in asthma and has serious implications that need to be addressed urgently. The use of dexmedetomidine in acute asthma and lung protection has been reported, but the exact mechanism is unclear. Objective: To investigate the effectiveness and mechanisms associated with dexmedetomidine in airway hyperresponsiveness. Methods: Forty BALB/c female mice were randomly divided into ve groups: group K (blank group), group A (asthma group), group HD (asthma + dexmedetomidine treatment group), group TH (asthma + yohimbine group) and group HT (asthma + dexmedetomidine + yohimbine group), and the airway resistance of group K, group A and group HD were analysed by invasive airway resistance assay, ELISA assay, immunohistochemistry and q-PCR, respectively. Airway resistance; IL-4 and IgE levels in serum and BLAF; and IL-4, IL-13, Muc5AC, NFκB, TLR2, TLR4 and TSLP1 protein levels in lung tissues of the 5 groups were analysed by invasive airway resistance assay, ELISA, immunohistochemistry and qPCR. Results: Compared with group A, there were statistical differences in airway resistance (P < 0.05); LIL-4 and IgE (P < 0.05) in serum and BLAF; and Muc5AC, TLR4 and NFκB protein contents (P < 0.05) in lung tissues in the HD group. Conclusion: 1. Dexmedetomidine can attenuate airway hyperresponsiveness in the OVA asthma model; 2. Dexmedetomidine reduced the production of IL-4 and IgE by down-regulating the TLR4/NF-κB signaling pathway, thereby reducing the lung inammatory response and airway hyperresponsiveness in the OVA-induced asthma model.

while some clinical patients with underlying allergies, unknowingly using some anesthetic drugs or uids can cause increased airway pressure. Airway hyperreactivity is a clinical manifestation that is insidious and unpredictable. In the perioperative period, when triggered by speci c factors, there is a tendency for airway spasm, sputum (and airway narrowing) to cause mechanical ventilation di culties leading to hypoxaemia and hypotension. In severe cases, a "silent lung" may develop, which can be life-threatening.
Asthma is a serious global health problem [6] and airway hyperresponsiveness is not only a major clinical manifestation of asthma, but also of other diseases, making it more widespread than asthma. In general, preoperative use of β2 agonists can be used as treatment or prevention for asthma patients, but there are no good preventive measures for patients with other potential airway hyperresponsiveness in the perioperative period, especially during peri-anaesthesia. In order to effectively protect patients' total airway safety and life safety during the perioperative period, it is an important issue to identify the mechanism of airway hyperreactivity and to provide effective preventive and therapeutic measures.
Dexmedetomidine is a highly selective α2-adrenergic agonist with sedative, analgesic and anxiolytic effects, and is one of the most widely used drugs in clinical anesthesia. The protective effect of dexmedetomidine on lung injury has been studied extensively [7,8] . In addition, the immunomodulatory effects of dexmedetomidine have been noted [9] . Studies have shown that dexmedetomidine affects immunity by modulating speci c Th2 helper cells [10,11] , which in turn are major players in asthma, mediating in ammatory responses and airway hyperreactivity through the production of type 2 cytokines (IL-4, IL-13, thymic stromal lymphopoietin (TSLP), etc.) [11] ; dexmedetomidine reduces in ammatory factor secretion by attenuating NFκB signalling pathway phosphorylation in monocytes and macrophages [12] . Meanwhile Hamida Hammad [11] noted that most allergens and many air pollutants trigger the NFκB signalling pathway to promote Th2 development by activating toll-like receptors (TLRs), among others, activating innate immune cells and subsequently producing many mediators that contribute to airway in ammation.Martijn L. Manson [13] found that it was IL-13 and IL- 4 We therefore hypothesise that dexmedetomidine reduces IL4/13 by downregulating the TLR4/NF-κB signalling pathway to alleviate airway hyperresponsiveness in OVA mice, thus achieving prevention of perioperative airway hyperresponsiveness.

Materials And Methods
Forty female BALB/c mice, 5-7 weeks old, were purchased from the Experimental Animal Centre of the Hospital of Orthopaedic Surgery, Chinese Academy of Medical Sciences and housed in the SPF Animal Centre of the Hospital of Orthopaedic Surgery, Chinese Academy of Medical Sciences. The mice were fed with sterilized chow and drinking water, and after one week of acclimatization in the animal house, a mouse airway hyperresponsiveness model was started.They were also treated and housed in groups according to the different interventions.
2.1 OVA sensitisation protocol and dexmedetomidine treatment 2.1.1 Asthma model: BALB/c mice, sensitised by intraperitoneal injection of ovalbumin (OVA) sensitising solution, were nebulised using a PRONEB compression nebuliser during the excitation period, and the model was successfully established for a total of 55 days.

Evaluation of successful asthma model establishment
Airway responsiveness monitoring: 48h after the last nebulised excitation, the airway responsiveness of BALB/c mice was measured using an American invasive small animal lung function meter. Different concentrations of Methacholine (Mch,Sigma) were nebulised and the Enhanced Pause (Penh) values were measured at the corresponding concentrations. The Penh value at normal saline (NS) inhalation was used as the base value and the airway responsiveness was evaluated using the maximum Penh/NS% and PC100 and compared with the control group.

Dexmedetomidine treatment protocol and grouping
After successful establishment of asthma, the mice were divided into 3 groups: Asthma group (A), Dexmedetomidine(Jiangsu Hengrui Pharmaceutical Co.)group(HDgroup), Yohimbine (Sigma)group(TH group) and Dexmedetomidine + Yohimbine group (HT group); group A was given the same dose of saline; group HD, a single intraperitoneal injection of dexmedetomidine 25ug/kg for 5 days; group TH, an intraperitoneal injection of yohimbine 1mg/kg for 5 days; in the HT group, a single intraperitoneal dose of dexmedetomidine 25ug/kg + yohimbine 1mg/kg for 5 days.

Collection of peripheral blood
Peripheral blood was collected within 24 hours of AHR stimulation. Mice were anaesthetised with Sevo urane (Baxter Healthcare Corporation) per mouse and blood was collected from the tail vein after anaesthesia. 50-100ul of blood was collected and left at room temperature for 1 to 2 hours, centrifuged at 3000r/min for 10 minutes and the serum was separated and stored at -20℃ for ELISA testing.

Bronchoalveolar lavage uid(BALF)
After collecting blood specimens from mice under anaesthesia, the mice were xed and disinfected, the trachea was surgically exposed, a small incision was made, a sterile 22G indwelling needle was inserted ( xed to avoid damaging the lung too deeply), the left lung was immediately ligated, 0.5ml of 0.9% NaCl solution at 4℃was slowly injected into the right lung with a syringe each time, and then slowly withdrawn. The total recovery was about 1.2 ml (80-90% recovery). The lysate was ltered through gauze and the resulting BALF was centrifuged at 2000 r/min for 10 minutes at 4℃. After centrifugation, the cells were resuspended in PBS and then stained with Giemsa stain, and the total number of cells and cell sorting were counted under a light microscope; the supernatant was separated and stored at -20℃for ELISA.

Lungs.
After obtaining the BALF, the catheter was removed and the lung tissue was quickly freed and separated.
The middle lobe of the right lung was xed in para n, sectioned, and stained for HE and immunohistochemistry. The fresh left lung was frozen at -80°C and subjected to Real-time PCR.(Lung specimens were collected within 24 hours of the end of the last nebulisation stimulation and intervention. Mice were rst anaesthetised by inhalation for 1 minute in a glass cylinder of ether, then their heads and limbs were xed to the table in a "large" position, the lungs and heart were exposed by opening the chest, the left lung was quickly ligated with a thin string, the left lung tissue was cut out and immediately packed into DEPC water-treated EP tubes and stored in a -80°C freezer for PCR. The lung tissue was then intubated through the right ventricle and rinsed by rapid perfusion with saline at 4°C until the lung lobes turned white. Finally, the lungs were embedded in para n wax and sliced to a thickness of approximately 4 µm. 6 sections of each specimen were selected for HE staining and immunohistochemical staining. Six sections of each specimen were selected for HE staining and immunohistochemical staining. In ammation was scored by observing the HE-stained lung changes in each group of mice under a light microscope using a high-powered eld of view (40x), and was divided into 0-4 points according to the degree of in ammatory cell in ltration around the bronchioles: 0, normal bronchioles with no in ammatory cell in ltration; 1, bronchioles with less than 1 layer of in ammatory cell in ltration; 2, bronchioles with 1 layer of in ammatory cell in ltration; 3, bronchioles with 2-4 layers of in ammatory cell in ltration; 4, bronchioles with more than 4 layers of in ammatory cell in ltration. in ltrates ne bronchi; 4 points, more than 4 layers of in ammatory cells in ltrate ne bronchi.
The left lung of the mice was soaked in 4% neutral formaldehyde, and the soaking solution was replaced with 30% sucrose water after 12h-24h of soaking, and the lung tissue continued to be soaked at room temperature for 12h-24h. After staining, the peribronchial cupped cell hyperplasia was observed microscopically and scored: the bronchi of each group of mice were observed under a high magni cation light microscope (10×40), and the degree of cupped cell hyperplasia was mainly evaluated microscopically. The score of cupped cell hyperplasia was 0-4 according to the proportion of cupped cells in the peritubular area of the bronchial wall: 0, no cupped cells in the bronchial epithelium; 1, less than 25% of the peritubular area of the bronchial wall; 2, 25%-50% of the peritubular area of the bronchial wall; 3, 51%-75% of the peritubular area of the bronchial wall; 4, cupped cells in the peritubular area of the bronchial wall The proportion of cup-shaped cells in the peritubular area of the bronchial wall is > 75%.

ELISA
The mouse serum and BALF supernatant stored at -20°C were removed and the IL-4 and IgE levels in the BALF uid and peripheral blood were measured in strict accordance with the ELISA kit instructions. At the end of the experiment, the Ct value of each PCR measurement was recorded and averaged three times, using β-action as the internal reference. The data were processed according to the 2-△△ Ct method. △ Ct = mean ± standard deviation of Ct values for (target gene -reference gene); △ △ Ct = mean ± standard deviation of △ Ct for (sample to be tested -reference sample); mean ± standard deviation of initial template amount for relative samples = (2 -△ △Ct). Relative expression of target gene (RQ value) = 2-△△ Ct

2.4Statistical analysis
In this study, Imege J was used for image processing.The data were analysed using GraphPad Prism 8.0 software. One-way ANOVA was used for comparison when the means of each group met the chi-square; when the means of each group did not meet the chi-square, the rank sum test was used. A multivariate − 3 ANOVA was used for comparison between groups. When P < 0.05, the difference is statistically signi cant, and when P < 0.01, the difference is statistically signi cant.

Results
3.1 Increased airway in ammatory response and presence of airway hyperresponsiveness in the OVA asthmatic BALB/c female mouse model.
After the OVA-induced allergic asthma BALB/c female mouse model was established, airway responses were stimulated by incremental doses of acetylcholine, and airway resistance was measured (as shown in Fig. 3-A), which was statistically different in group A compared to group K at 6h, 12h, 24h and 46h after OVA induction. Further observation of pathological tissues (HE and PAS staining, found in Fig. 3-B and C), compared with group K, a large number of in ammatory cells were seen in the ne bronchial wall and interstitial lung stroma in group A, and there was signi cant destruction of lung tissue; also visible was cupular cell hyperplasia and increased mucus secretion, which was also statistically signi cant after scoring analysis ( Figure D). IL-4 and IgE were also measured in blood (Blood) and bronchoalveolar lavage uid (BLAF) by ELISA ( Fig. 3-E, F, G), and were statistically different in group A compared to group K. To later investigate the underlying mechanism, we measured Muc5Ac, NFκB, and TLR4 in lung tissue by immunohistochemistry (Fig. 3-H), which were signi cantly higher in lung tissue in group A compared with group K (Figure I, P < 0.001). IL-4 was elevated in lung tissue, blood, and BALF uid after OVA exposure; IgE was elevated in blood IL-4 was elevated in lung tissue, blood and BALF uid; IgE was elevated in blood and BALF uid; Muc5AC, NFκB and TLR4 were elevated in lung tissue.
After ve consecutive days of treatment with dexmedetomidine 25ug/kg by intraperitoneal injection respectively, invasive airway resistance was measured by the same method in the HD group of OVA asthma mouse model. Compared with group A, airway resistance in the HD group was different at 48h, interestingly, airway resistance was signi cantly reduced at 12h, but P = 0.05, no statistical difference, but there was a statistical difference from 12h-48h (Fig. 2-A).

Dexmedetomidine improves lung pathology after OVA exposure
The total number of eosinophils and neutrophils in the lungs of the HD group was found to be signi cantly reduced by Giemsa staining and counting of eosinophils and neutrophils under electron microscopy ( Fig. 5-A) compared to group A. There was a statistically signi cant difference (Fig. 5-B). Further observation of pathological tissues (HE and PAS staining, found in Figs. <link rid=" g3">5</link>-C and 5-E) showed a reduction in in ammatory cell in ltration in the ne bronchial walls and interstitial lung stroma and a signi cant reduction in lung tissue destruction in group K compared to group A. After scoring the two groups by multiple hypothesis testing (Tukey HSD post hoc test (Fig. 5-D), HD was lower than the mean of A. The difference between the two groups was − 13.791 (− 19.704 --7.877), a statistically signi cant difference (P < 0.001); meanwhile, reduced mucus secretion was found after PAS staining, and after scoring analysis (Tukey HSD post hoc test) (Fig. 5-F), cupulocytosis was diminished and the HD group was below the mean of A, with a difference of -20.488 between the two groups (-27.192 -− 13.785), with a statistically signi cant difference (P < 0.001).
3.4 Dexmedetomidine inhibits the expression of markers of allergic airway in ammation following OVA exposure.
In OVA-exposed mice treated with dexmedetomidine, IL-4 levels in peripheral blood and BLAF uid were measured by ELISA and analysed by t-test, and were signi cantly reduced in the HD group compared to group A. The difference was 0.018 (0.002-0.033) ,(P = 0.036) in blood (Fig. 6-A) and in BLAF uid (Fig. 6-B) 0.076 (0.05-0.102), the difference was statistically signi cant ( P = 0.001). In contrast, there was no statistical difference between the TH group (group with α2-blocker, yohimbine) and group A (Figure-6C), and the IL-4 protein variables in lung tissue were detected by qPCR ( Fig. 6-E), which was also statistically different. Subsequently, we detected the immunomarker IgE in blood and BALF uid by ELISA (Figure-6D), and there was a statistical difference in both HD groups compared to group A. It can be seen that dexmedetomidine reduces the markers of allergic airway in ammation IL-4 and IgE after OVA exposure.
3.5 Dexmedetomidine inhibits the in ammatory response through TLR4/NFκB2 channels, but not TLR2; it can affect IL-4 but not IL-13.
To clarify the speci c mechanism by which dexmedetomidine attenuates the in ammatory response after OVA exposure, it was previously reported that the TLR/NFκB channel is one of the important channels causing the in ammatory markers in the allergic lung. It is known from previous studies that TLR4/NFκB is signi cantly elevated after OVA exposure. Therefore, we once again examined Mu5AC, TLR4 and NFκB in the HD group and the A and HT groups (dexmedetomidine + yohimbine group) by immunohistochemistry. Compared with the A and HT groups, Mu5AC, TLR4 and NFκB were signi cantly lower and statistically different in the HD group; compared with the A group, there was no statistical difference in the HT group. To clarify the speci c changes in proteins in the A and HD groups, the protein content of IL-4, IL-13, Muc5AC, NFκB, TLR2, TLR4 and TSLP1 in lung tissue was measured by q-PCR.
There were no signi cant changes in IL-13 and NFκB in lung tissue in the HD group compared to group A. It is possible that dexmedetomidine attenuated the in ammatory response independently of TLR2/NFκB and had no effect on IL-13.

Discussion
Asthma is a disease that affects health worldwide. Airway hyperresponsiveness is not only the main clinical manifestation and pathophysiological feature of asthma, but is also stimulated by certain speci c substances in the environment. It can cause hypoxaemia and even asphyxia in the perioperative period. Current research suggests that the mechanisms involved in airway hyperresponsiveness are neurosensitisation, in ammatory bursts, activation of the immune response, epithelial cell chemotaxis, cupped cell proliferation and increased mucus secretion [11,[14][15][16] .
Hamida Hammad [11] and others published in cell journal concluded that most allergens and many air pollutants trigger epithelial cell nuclear factor kB (NF-kB) production by activating toll-like receptors (TLRs) or protease-activated receptors (PARs), leading to activation of dendritic cells (DCs) that migrate to priming nodes to promote Th2 development and activate the recruitment of innate immune cells to the airways, which subsequently produce many of the mediators that contribute to airway in ammation.
Related studies have reported that neutrophils amplify recruitment and in ammatory responses in neutrophilic asthma by stimulating airway epithelial cells to activate the TLR4/NF-κB pathway and secrete chemokines [17] ; improvement of lipopolysaccharide-induced injury, in ammation and MUC5AC expression in bronchial epithelial cells by inhibiting TLR4/NF-κB signalling has also been reported [18] . In contrast, MUC5AC is one of the main proteins of airway hyperresponsiveness, causing more mucous sputum secretion by cupped cells [19,20] . It can be seen that TLR4/NF-κB signalling pathway has an important role in airway in ammatory response and airway hyperresponsiveness. In our study, we found that TLR4/NF-κB was signi cantly increased after OVA exposure, as were the in ammatory factor IL-4 and the antigen-presenting agent IgE. This is consistent with the relevant studies reported above.
Dexmedetomidine is a highly selective α2 adrenergic agonist (α2:α1 selectivity of 1600:1). α2 receptors are G protein-coupled receptors with norepinephrine and epinephrine as ligands. By binding to ligands it leads to Gi/G0 protein coupling, attenuates adenylyl cyclase, inhibits voltage-gated Ca 2+ channels and promotes K + inward ow thereby hyperpolarising cells. α2 receptors are widely expressed in the central and peripheral nervous system as well as on other cells and tissues such as leukocytes and endothelial cells [9,21] .
Currently, dexmedetomidine is one of the most widely used anaesthetic agents in the perioperative period and in the ICU. Dexmedetomidine has been widely reported to reduce ventilator-induced lung injury by activating the ERK1/2 pathway [7] and to reduce non-ventilator-related lung injury by modulating the immune phenotype of macrophages [22] ; it can also improve endotoxin-induced acute lung injury in vivo by maintaining mitochondrial dynamic homeostasis through the HIF-1a/HO-1 signalling pathway [8] ; it has also been related demonstrated its effectiveness in chronic obstructive pulmonary disease [23] . In the present study, dexmedetomidine was shown to attenuate airway resistance in an OVA-induced asthma model, and its protective effect on the lung was again con rmed by sectioning and staining of lung tissue.
In ammatory responses and immune responses are key to the development and progression of airway hyperresponsiveness. The main speci c cytokines associated with in ammation are IL-4, IL-5 and IL-13 secreted by Th2 cells; the main protein associated with the immune response allergen-speci c B cells synthesise immunoglobulin E (IgE) [9,14] . In the current study, IL-4 and IgE were measured in each group by ELISA and dexmedetomidine was found to moderate lung in ammation by modulating IL-4 and IgE. However, analysis of IL-13 in each group by q-PCR revealed that dexmedetomidine did not have a statistically signi cant modulating effect on IL-13. We know that both IL-4 and IL-13 are secreted by Th2 cells, however, IL-4 production is dependent on calcium-regulated neurophosphatase, whereas IL-13 production is only partially dependent on calcium-regulated neurophosphatase [16 ] . Calcium-regulated neurophosphatase is positively correlated with intracellular Ca 2+ [24] , and dexmedetomidine is an α2 receptor agonist, and α2 receptor agonism inhibits voltage-gated Ca 2+ channels [9] . Therefore, we conjecture that dexmedetomidine may reduce intracellular Ca 2+ concentration by inhibiting Ca 2+ inward ow, thereby reducing the amount of calcium-regulated neurophosphatase and may be partly responsible for the lower IL-4 production. Unfortunately, we did not consider that IL-4 and IL-13 are not produced in the same direction at the beginning of our project, so we did not design the neural calcium-regulated neurophosphatase blockade.
Toll-like receptors (TLRs) are a subfamily of pattern recognition receptors that recognise invading pathogens and endogenous noxious stimuli to induce innate and adaptive immune responses. Among the TLRs, TLR4 is the major lipopolysaccharide (LPS) receptor. When expressed on the cell membrane TLR4 activates downstream mediators, including the transcription factor nuclear factor (NF)-κB. TLR4/NF-κB can increase cytokines (e.g. tumour necrosis factor (TNF)-α, interleukin (IL)-1β, IL-6), chemokines, enzymes, reactive oxygen species [25] . Several studies have demonstrated that the in ammatory response can be suppressed by inhibiting the TLR4/NF-κB signalling pathway [25,26] . In anti-in ammatory lung injury [27] and protection against ischemia-reperfusion injury [28] , dexmedetomidine has been reported to attenuate the in ammatory response by inhibiting the TLR4/NF-κB signalling pathway. In this study, the expression of TLR4/NF-κB in different groups was analysed by immunohistochemistry and q-PCR, which not only rea rmed the pulmonary protective effect of dexmedetomidine, but also con rmed the anti-in ammatory effect and attenuated airway hyperreactivity in the airways. In immunohistochemistry, the effect of dexmedetomidine was attenuated in the yohimbine group, showing that α2 receptors are involved in the inhibition of TLR4/NF-κB signalling pathway generation. However, no effect on TLR2 was seen, and related reports suggest that nebulised lidocaine inhibits airway smooth muscle cell proliferation and migration by downregulating TLR2 to improve allergic airway in ammation [29] and blocking the TLR2/NF-κB/NFATc1 signalling pathway [30] . According to the literature, activation and correct assembly of TLR4 with upstream differentiation protein 2 (MD-2) requires relatively stable concentrations of cations of substances such as Ca 2+ and Mg 2+ [25,31] ,while TLR2 is upstream of Pam3CSk4 [29,33] . We speculate that perhaps dexmedetomidine affects MD2 and thus TLR4/NF-κB expression.
Muc5AC protein is the major gel-forming mucin in airway secretions. Increased airway mucus contributes to airway obstruction in respiratory diseases such as asthma and chronic obstructive pulmonary disease. muc5AC gene and protein expression is regulated by cytokines such as IL-4 and IL-17 [32] . In this experiment, Muc5AC protein levels were measured by q-PCR in the asthma and dexmedetomidine treated groups and were found to be suppressed by dexmedetomidine in line with changes in IL-4.
Earlier, according to DS Ferreira et al [34] , thymic stromal lymphopoietin1 (TSLP1) was reported to be highly expressed in the airways of acute lethal asthma, together with TLR4. Recently, it was reported that eosinophils are elevated in patients with severe asthma and can activate lung epithelial cells to produce IL-33 and TSLP [35] . TSLP 1 is frequently expressed in severe asthma. In this study, TSLP1 expression was lower in the dexmedetomidine group than in the asthma group as determined by q-PCR. The speci c intrinsic regulatory mechanisms were not addressed in this study, but only correlations were seen, which may provide new therapeutic ideas for future patients with severe or fatal asthma.
In summary, dexmedetomidine could downregulate the expression of TLR4/NF-κB signalling pathway by inhibiting Ca 2+ inward ow, which in turn affected the secretion of cytokine IL-4 by T2h cells and immunoglobulin E by B cells; the reduction of cytokine IL-4 led to a decrease in Muc5AC expression. The speci c mechanism of dexmedetomidine regulation of TSLP is not clear from this study. The datasets used and/or analyzed during the current study are available from the corresponding author on reasonable request.

Competing interests
The authors have declared that there is no con ict of interest.  The present study suggests Dexmedetomidine attenuates in ammatory cytokines and airway hyperresponsiveness in models of asthma, and there are possible mechanisms by which. 1.
dexmedetomidine downregulates the TRL4/NFκB signalling pathway by inhibiting Ca2+ inward ow; 2. down-regulation of TRL4/NFκB reduces stimulation of TH2 cells, thereby decreasing IL4 and IGE secretion and consequently Muc5AC protein expression; 3.The decrease in TSLP expression after dexmedetomidine application may be related to the downregulation of the TRL4/NFκB signalling pathway and possibly to attenuated epithelial cell stimulation; 4.The effect of dexmedetomidine on TH2 cells may have the involvement of calcium-regulated neurophosphatase.

Figure 2
Experimental process diagram. A shows that the airway resistance was signi cantly higher in the asthma group than in the blank group after passing the modelling, and all were statistically different after 6h; B, shows the HE staining results.C, shows PAS staining results. By B C D, we know that after the asthma model, there was severe structural destruction of the lung, increased in ammatory in ltration, increased sputum, and cupped cell hyperplasia. Based on E F G, we know that IL-4 and IGE in both peripheral serum and BALF were increased in the asthma model. His the result of immunohistochemistry of lung tissue. According to H and I, Muc5AC, TRL4/NFκB were increased in lung tissues of the asthma model. The graph shows that airway resistance decreased signi cantly at all time periods after dexmedetomidine treatment compared to the asthma group. However, interestingly no statistical difference was seen at the 12h time point (P=0.05) Figure 5 A shows that the total number of eosinophils and neutrophils was signi cantly reduced after dexmedetomidine treatment by Kimsa staining of a portion of BALFA uid. B C D shows retarded structural destruction of the lung, diminished cuprocyte chemotaxis and reduced mucus after dexmedetomidine treatment. Figure 6 Graphs A, B and D show a decrease in IL-4 and ige in serum and BALF after dexmedetomidine treatment compared to the asthma group (group A). There was also a decrease in IL-4 in lung tissue as veri ed by pcr testing, as shown in E. Yohimbine was also used as a drug control group and was found to have no signi cant effect on IL-4 in serum and BALF, which is shown in C.

Figure 7
A shown, is the expression of Muc5AC, TRL4/NFκB in immunohistochemistry in the lung tissue of each group in response to group A, HD and HT. By B we can know that the expression of Muc5AC, TRL4/NFκB all decreased after dexmedetomidine treatment and this effect was affected by yohimbine.