| Objective:To investigate therapeutic potential and mechanism of immunoglobulin Yolk(IgY) against tumour necrosis factor-alpha (TNF-α) and interleukin-1beta (IL-1β) ontreating guinea pigs with allergic rhinitis induced by ovalbumin (OVA) andalleviating inflammation of the lung tissue in guinea pigs with allergic rhinitisinduced by OVA.Methods:1. The preparation of IgY.2.To establish allergic rhinitis model in guinea pigs using ovalbumin viaintraperitoneal injection for sensitization and intranasal instillation for challenge.3. To identify allergic rhinitis model in guinea pigs with allergic rhinitis usingovalbumin via intradermal injection.4. Hartley guinea pigs were randomly divided into seven groups: the controlgroup (Group C), the allergic rhinitis model group (Group M), the nonspecific IgYtreatment group (Group Z1), the0.1%anti-TNF-α IgY treatment group (Group Z2),the0.1%anti-IL-1β IgY treatment group (Group Z3), the0.1%anti-TNF-α and IL-1βIgY treatment group (Group Z4), and the fluticasone propionate treatment group(Group Z5). The results were observed at2h,4h and8h after the last treatment.5. The challenge was performed via intranasal with physiological saline in thecontrol group (group C), via intranasal with ovalbumin in the allergic rhinitis modelgroup (Group M) and via intranasal with ovalbumin after instilling different IgYantibodies and fluticasone propionate suspension in the nonspecific IgY treatmentgroup (Group Z1), the0.1%anti-TNF-α IgY treatment group (Group Z2), the0.1%anti-IL-1β IgY treatment group (Group Z3), the0.1%anti-TNF-α and IL-1β IgYtreatment group (Group Z4) and the fluticasone propionate treatment group (Group Z5). Within30min after OVA challenge the number and amount of sneezing, noserubbing and rhinorrhea were counted and scored in every group.6. The guinea pigs were anesthetized using10%chloral hydrate at2h,4h and8h after the last treatment and ovalbumin challenge, the nasal cavity was lavaged byinstilling PBS buffer and the nasal lavage fluid (NLF) was collected, as well as theNLF was centrifuged and it was separated into supernatant and precipitated cells.7. The guinea pigs were anesthetized using10%chloral hydrate at2h,4h and8h after the last treatment and ovalbumin challenge, the blood was gotten by cardiacpuncture and was anticoagulated, the blood smear was prepared, as well as the bloodwas centrifuged and the plasma was separated.8. The guinea pigs were anesthetized using10%chloral hydrate at2h,4h and8h after the last treatment and ovalbumin challenge. After the blood was gotten, theguinea pigs were put to death and the bronchoalveolar lavage was performed usingPBS buffer, the bronchoalveolar lavage fluid (BALF) was centrifuged, as well as theprecipitated cells were smeared.9. The guinea pigs were anesthetized using10%chloral hydrate at2h,4h and8h after the last treatment and ovalbumin challenge. After the bronchoalveolar lavagewas performed, the nasal mucosa and the left lung of guinea pigs were separated,immersed and fixed using4%paraformaldehyde, embedded in paraffin, as well as thehistologic section was prepared.10. The levels of cytokines (e.g. TNF-α, IL-1β, IL-5, IL-9, IL-13, IL-18, IL-22,IL-33, TGF-β) and OVA-specific IgE in plasma and nasal lavage fluid (NLF)supernatant were determined using the sandwich enzyme-linked immunosorbentassay (ELISA) method.11. The smeared cells in the peripheral blood, nasal lavage fluid andbronchoalveolar lavage fluid were stained using Wright`s solution, and the percentageof eosinophils, neutrophils, lymphocytes and macrophages was calculated.12. The histologic sections of the nasal mucosa and lung tiuess were stainedusing hematoxylin-eosin staining (HE). The inflammatory histopathology andinfiltrated eosinophils in the nasal mucosa and lung tiuess sections were observed andcalculated 13. The TNF-α, IL-1β, IL-5and IL-33expressions were observed usingimmunohistochemistry staining in the nasal mucosa and lung tiuess sections.14. All data were analyzed using SPSS19.0statistical software package.Results:1. The success rate on establishing allergic rhinitis model of guinea pigs inducedby ovalbumin via intraperitoneal injection for sensitization and intranasal instillationfor challenge was96.63%(172guinea pigs/178guinea pigs) and the mortality was69.85%(183guinea pigs/262guinea pigs).2. The allergy symptoms and number of sneezing, nose rubbing and rhinorrheain guinea pigs with allergic rhinitis followed OVA stimulation within30min wereobviously alleviated and decreased in the Z2, Z3, Z4and Z5groups compared to the Mgroup (P<0.05). The guinea pigs in group C occasionally had mild nose rubbing.3. There was little lymphocyte infiltration in the nasal mucosa and lung tissuein group C. The allergic inflammation pathologic response, such as the eosinophil andlymphocyte infiltration, submucosa edema, and the integrity of mucosal epithelial celllayer, was alleviated in the Z4and Z5groups compared to the M and Z1groups,meanwhile it was lighter than that in the Z2and Z3groups, particular at8h in thenasal mucosa. The allergic inflammation pathologic response, such as the eosinophil,neutrophil and lymphocyte infiltration, bronchial mucosa epithelial cell falling off,pulmonary interstitial edema, alveolar ducts damage, thickening and breakage ofalveoli septum and thickening of bronchial smooth muscle, was alleviated in the Z4and Z5groups compared to the M and Z1groups, meanwhile it was lighter than that inthe Z2and Z3groups, particular at8h in the lung tissue.4. The eosinophils percentage of decrease in the nasal mucosa and lung tissuewas in turn the C group <the Z5group <the Z4group <the Z3group <the Z2group <the Z1and M groups from2h to8h, and the eosinophils percentage was significantlydecrease in the Z4and Z5groups compared to the M and Z1groups (P<0.05).5. The expressions of TNF-α, IL-1β, IL-5and IL-33mainly were in the epithelialcells, and gland cells of intrinsic and submucosal layers in the nasal mucosa, as wellas in the gland cells of alveoli and alveolar septum and epithelial cells of bronchial mucosa in the lung tissue. The expression quantity of IL-1β and IL-33was obviouslydecrease in the Z4and Z5groups compared to the M, Z1,Z2and Z3groups. These wereno obvious difference between the M group and the Z1group, and the Z4group andthe Z5group.6. The eosinophil in the Z2, Z3and Z5groups at2h and the Z4group at8h in theperipheral blood was significantly lower than that in the M group (P<0.05). Theeosinophil was significantly decreased from2h to8h in the Z2, Z3,Z4and Z5groupscompared to the M and Z1groups, the neutrophil was significantly decreased in groupZ5at2h, in the Z3, Z4and Z5groups at4h, and in the Z3and Z5groups at8hcompared to the M group, and the lymphocyte was significantly decreased in the Z4and Z5groups at2h, in group Z4at4h and in the Z3and Z5groups at8h compared tothe M group in the NLF (P<0.05). The eosinophil was significantly decreased in theZ2,Z3, Z4and Z5groups at2h, in the Z3, Z4and Z5groups at4h and8h compared tothe M group, the neutrophil was significantly decreased in the Z2, Z3, Z4and Z5groups at2h and in group Z4at8h compared to the M group, and the lymphocytewas significantly decreased in the Z1, Z3, Z4and Z5groups at2h, in the Z3and Z5groups at4h and in group Z5at8h compared to the M group in the BALF (P<0.05).7. The levels of IL-1β, TNF-α, IL-5, IL-9, IL-33and OVA specific-IgE weresignificantly decreased in the Z2, Z3, Z4and Z5groups compared to the M and Z1groups, the level of IL-13was significantly decreased in the Z3, Z4and Z5groupscompared to the M, Z1and Z2groups and the levels of IL-18, TGF-β and IL-22weresignificantly decreased in the Z2, Z3, Z4and Z5groups compared to the M group at2hin the NLF (P<0.05). The levels of IL-1β, IL-5, IL-9, IL-33and OVA specific-IgEwere significantly decreased in the Z2, Z3, Z4and Z5groups compared to the M andZ1groups, the level of IL-13was significantly decreased in the Z3, Z4and Z5groupscompared to the M and Z2groups, the level of TNF-α was significantly decreased inthe Z2,Z3, Z4and Z5groups compared to the M group, the level of TGF-β wassignificantly decreased in the Z3, Z4and Z5groups compared to the M group, and thelevel of IL-22was significantly decreased in the Z4and Z5groups compared to the Mgroup at4h in the NLF (P<0.05). The levels of IL-1β, TNF-α, IL-5, IL-9and IL-33were significantly decreased in the Z2, Z3, Z4and Z5groups compared to the M group, the levels of IL-13and IL-22were significantly decreased in the Z3, Z4and Z5groupscompared to the M group, the level of TGF-βwas significantly decreased in the Z2, Z4and Z5groups compared to the M group, the level of OVA specific-IgE wassignificantly decreased in the Z4group compared to the M group at8h in the NLF(P<0.05). The levels of IL-1β, IL-9, IL-22, TGF-β and OVA specific-IgE weresignificantly decreased in the Z2, Z3, Z4and Z5groups compared to the Z1group, thelevels of IL-5, IL-33and TNF-α were significantly decreased in the Z3, Z4and Z5groups compared to the Z1group and the level of IL-13was significantly decreased inthe Z5group was decreased compared to the Z1group at8h in the NLF (P<0.05).8. The levels of TNF-α, IL-13, TGF-β, IL-33and OVA specific-IgE weresignificantly decreased in the Z2, Z3, Z4and Z5groups compared to the M and Z1groups, the levels of IL-5and IL-9were significantly decreased in the Z2, Z3, Z4andZ5groups compared to the M group, the levels of IL-9, IL-13and TNF-α weresignificantly decreased in the Z1group compared to the M group, the level of IL-18was significantly decreased in the Z3, Z4and Z5groups compared to the M group andthe level of IL-1β was significantly decreased in the Z4and Z5groups compared to theM group at2h in the plasma (P<0.05). The level of IL-5in the Z3, Z4and Z5groups,the level of IL-9in the Z4group, the level of IL-18in the Z3and Z4groups and thelevels of IL-1β and IL-22in the Z4and Z5groups were significantly decreasedcompared to the Z1group at2h in the plasma (P<0.05). The levels of IL-5, IL-18,TGF-β, IL-22and IL-33were significantly decreased in the Z2, Z3, Z4and Z5groupscompared to the M and Z1groups, the level of IL-13in the Z1, Z2, Z3, Z4and Z5groups, the level of OVA specific-IgE in the Z2, Z3, Z4and Z5groups, the levels ofIL-1β and IL-9in the Z3, Z4and Z5groups and the level of TNF-α in the Z4groupwere significantly decreased compared to the M group at4h in the plasma (P<0.05).The level of OVA specific-IgE in the Z3, Z4and Z5groups, the level of IL-9in the Z3,Z4and Z5groups, the level of IL-13in the Z3and Z5groups, the level of IL-1β in theZ4and Z5groups and the level of TNF-α in the Z4group were significantly decreasedcompared to the Z1group at4h in the plasma (P<0.05). The levels of IL-5, TGF-βand IL-33were significantly decreased in the Z2, Z3, Z4and Z5groups compared tothe M and Z1groups, the level of OVA specific-IgE in the Z5group, the levels of IL- 1β and IL-9in the Z4and Z5groups, the level of IL-13in the Z1, Z2, Z3and Z4groups,the level of IL-18in the Z4group, the level of TNF-α in the Z2and Z5groups and thelevel of IL-22in the Z1, Z2, Z3, Z4and Z5groups were significantly decreasedcompared to the M group at8h in the plasma (P<0.05). The level of OVA specific-IgE in the Z5group, the level of IL-9in the Z4and Z5groups, the levels of TNF-α andIL-18in the Z2, Z3, Z4and Z5groups and the level of IL-22in the Z2group weresignificantly decreased compared to the Z1group at8h in the plasma (P<0.05).9. The level of TNF-α was positively correlated with that of IL-1β (r=0.623,P<0.01), IL-5(r=0.676, P<0.01), IgE (r=0.703, P<0.01), IL-9(r=0.634, P<0.01),IL-13(r=0.541, P<0.01), IL-18(r=0.703, P<0.01), TGF-β (r=0.779, P<0.01), IL-22(r=0.324, P<0.01) and IL-33(r=0.723, P<0.01), and the level of IL-1β waspositively correlated with that of IL-5(r=0.677, P<0.01), IgE (r=0.723, P<0.01),IL-9(r=0.745, P<0.01), IL-13(r=0.576, P<0.01), IL-18(r=0.558, P<0.01), TGF-β(r=0.728, P<0.01), IL-22(r=0.515, P<0.01) and IL-33(r=0.750, P<0.01) in theplasma. The level of TNF-α was positively correlated with that of IL-1β (r=0.761,P<0.01), IL-5(r=0.619, P<0.01), IgE (r=0.657, P<0.01), IL-9(r=0.736, P<0.01),IL-13(r=0.549, P<0.01), TGF-β (r=0.625, P<0.01), IL-22(r=0.704, P<0.01) andIL-33(r=0.691, P<0.01), and the level of IL-1βwas positively correlated with that ofTNF-α (r=0.761, P<0.01), IL-5(r=0.754, P<0.01), IgE (r=0.821, P<0.01), IL-9(r=0.877, P<0.01), IL-13(r=0.583, P<0.010, IL-18(r=0.402, P<0.01), TGF-β (r=0.721, P<0.01), IL-22(r=0.719, P<0.01), and IL-33(r=0.779, P<0.01) in the NLF.Conclusion:1. The allergic rhinitis model in guinea pigs is successfully established usingovalbumin and aluminium hydroxide via intraperitoneal injection for sensitizationand ovalbumin via nasal instillation for challenge.2. The pathological development of allergic inflammation pathological responsein the nasal mucosal and lung tissue is effectively inhibited using anti-TNF-α and IL-1β IgY via intranasal instillation in guinea pigs with allergic rhinitis induced by OVA,and the allergic inflammation is obviously alleviated. 3. The mechanism of anti-TNF-α and IL-1β IgY treating in guinea pigs withallergic rhinitis may be due to reduce the increase and infiltration of inflammationcells, such as eosinophils, lymphocytes and neutrophils, and inflammatory cytokines,such as IL-1β, TNF-α, IL-5, IL-9, IL-13, IL-22, IL-33and TGF-β1, and OVA specificIgE in the nasal mucosal and lung tissue and the peripheral blood and nasal lavagefluid (NLF) and bronchoalveolar lavage fluid (BALF).4. The occurrence and development of inflammation pathological response ofallergic bronchial asthma in guinea pigs with allergic rhinitis induced by OVA is asufficient proof that the allergic inflammation in the upper and lower airway is thesynchronization and consistency, and meanwhile the treatment also is consistent. |
PDF Full Text Request