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The S \u0026 P stomach pill (Puwei Pill, PWP) recipe pharmaceutical for the treatment of gastric ulcers (the Gastric Ulcer GU), the medication fewer lasting effect, the recurrence rate is low, with few side effects. Previously confirmed, PWP has a protective effect on a variety of experimental GU. Objective: Preparation of rat GU relapse model to study the role of the PWP anti GU and its recurrence. 2 visits PWP process of ulcer healing and recurrence of gastric mucosal inflammation and inflammatory factor nuclear transcription factor-κB (nuclear factor kappa-B, NF-κB), interleukin -8 (interleukins-8, IL-8), tumor necrosis factor-α (Tumor necrisis factor-α, TNF-α) protein expression from inflammation angle to reveal the PWP anti-GU and recurrence of mechanisms to provide a theoretical basis for further research and development of this drug. Method: the PWP anti GU recurrence of acetic acid irritating rat GU model, and then prepared by intraperitoneal injection of IL-1β GU relapse model, and then study the the PWP anti-chronic GU and its recurrence role and explore ulcer healing and relapse process PWP anti-inflammatory mechanism of gastric mucosal protection, designed to provide the theoretical basis for the ulcer index (ulcer index, UI) and the rate of recurrence of ulcers, gastric mucosa pathology and inflammatory cell density for further research and development of this drug, visits PWP anti GU and The recurrence role. 2. PWP anti-inflammatory mechanism of GU occurrence and recurrence process using immunohistochemistry (of Immunohistochemistry, IHC) method, and immunoblotting (Western blot) method, observation of NF-κB PWP inflammatory cytokines, IL-8, TNF- α protein expression using radioimmunoassay detection PWP plasma IL-8 and TNF-α levels, and to examine the the PWP GU occurrence and recurrence of anti-inflammatory mechanism. Results: (1) 1.1 PWP the PWP anti GU their recurrence role of the GU rats UI and the relapse rate with acetate modeling 16d, model no recurrence (MWR) set of UI was 30.15 ± 6.40, omeprazole (OMLZ ) group and the PWP group were 20.32 ± 6.80 and 7.75 ± 2.09, compared with MWR group has significant difference (P lt; 0.01); 92d, the normal group, sham group and MWR group showed no recurrence of ulcers; model of relapse (MR) group ulcer recurrence rate was 83.3%, while OMLZ group was 33.3% the the PWP group, 10.0%, compared with the MR group, there were significant differences (P lt; 0.01). The results show that the PWP has better anti GU their recurrence. 1. 2 PWP GU recurrence of stomach histopathology 16d, HE staining showed that the structural integrity of the gastric tissue of the normal group and sham group, mucosal glands arranged in order; MWR group visible ulcers, mucous membrane detachment or defect , mucosa glands normal structure disappeared, ulcers infringement mucosa full-thickness and deep muscle or deeper, was necrotic lesions muscularis fracture submucosa lymphocytes, monocytes and other inflammatory cell infiltration, ulcer edge mucosa glands interstitial congestion and edema the, glands expansion; OMLZ group and the PWP group of ulcer mucosal defects was significantly reduced, non-exudative necrosis, the partially visible the regenerative mucosa to mouth ulcers migration, the rest of ulcer healing regenerative mucosa than the surrounding mucosa thin myometrial defect width smaller, inflammatory cell infiltration fewer glands arrangement compared with the overall expansion of the relatively small number. 92d, the normal group and sham group gastric tissue structural integrity, no ulcers, no significant inflammatory cell infiltration; MWR group no ulcers, visible thinner regenerative mucosa, scar tissue under the new granulation tissue scar mucosal inflammatory cell infiltration; the MR group relapse ulcer edge visible residues regenerative mucosa and inflammatory cell infiltration more; the OMLZ group and the PWP group recurrent ulcer regenerative mucosa thicker, less inflammatory cell infiltration. GU relapse rat gastric tissue inflammatory cell density 16d 1.3 PWP normal group and sham operation group of chronic inflammatory cell density were 0.30 ± 0.03 and 0.39 ± 0.03, between the two there is no significant difference. MWR group was 41.11 ± 2.94, significantly higher than the sham group (P lt; 0.01), the inflammatory cells are involved in the generation of GU. And OMLZ group and the PWP group were 21.25 ± 1.81 and 22.58 ± 2.10, and MR group ratio decreased significantly (all P lt; 0.01). Show that the PWP can significantly reduce the infiltration of inflammatory cells in the ulcer process. 92d, inflammatory cell density of the normal group and sham group were 0.31 ± 0.03 and 0.41 ± 0.03, between the two without a significant difference. MWR group and the MR group were 15.79 ± 1.14 and 60.83 ± 3.26, were significantly increased compared with the sham group (all P lt; 0.01), GU relapse involving the change of inflammatory cells. OMLZ group and the PWP group were 30.51 ± 1.95 and 31.18 ± 2.49, compared with the MR group decreased significantly (P lt; 0.01). Show that the PWP can significantly reduce the infiltration of inflammatory cells in the process of GU recurrence. The inflammatory cell density and UI correlation analysis found, 16d time, chronic inflammatory cell density and UI into related (r = 0.902 P lt; 0.01); 92d when, inflammatory cell density and recurrence rate as being related (r = 0.611, P lt; 0.01). Indicating that the inflammatory response involved in GU recurrence, PWP anti-inflammatory response by inhibiting gastric tissue GU and its recurrence role. 2 PWP GU occurrence and recurrence of anti-inflammatory Mechanism 2.1 PWP GU and its recurrence rat NF-κB 2.1.1 cellular localization of the NF-κB positive cells: NF-κB-positive cells expressing cell cytoplasm and nucleus of two states, namely, NF-κB inactivation and activation status. In the normal group and sham group, NF-κB expression in the cytoplasm of epithelial cells of the gastric mucosa layer, causing mold section 16d, MWR group mucosal epithelial cell cytoplasm and nucleus expression, its nucleus was significantly increased The nucleus of the OMLZ group and the PWP group expression MWR group reduced. 92d when the mucosa at the nuclear expression of the MR group ulcer edge compared with the MWR group significantly increased interstitial inflammatory cell nuclei were strongly positive, mainly OMLZ group, PWP the Zeyi cytoplasmic expression. 2.1.2 NF-κB-positive cells expressing the results of immunohistochemistry ⑴ 16d, the normal group, sham operation group, NF-κB positive cells were very small, 0.41 ± 0.39, 0.58 ± 0.31, respectively, the two groups had no significant difference (P gt; 0.05); compared with the normal group, MWR group significantly with the increase (11.75 ± 0.43) (P lt; 0.01), OMLZ group and the PWP group positive cell number respectively, 5.09 ± 0.16 and 5.23 ± 0.24, and MWR group compared the expression of positive cells were significantly lower (all P lt; 0.01). ⑵ 92d, the normal group, sham group and MWR positive cell counts were 0.49 ± 0.23,0.55 ± 0.33,1.40 ± 0.29, three showed no significant differences (all P gt; 0.05); the MR groups to 8.52 ± 0.95 MWR group significantly increased (P lt; 0.01); OMLZ group and the PWP group positive cell number, respectively, 4.68 ± 0.53 and 4.63 ± 0.32, with the MR group comparison is to reduce (all P lt; 0.01); OMLZ group and PWP group showed no significant sex difference (P gt; 0.05). Show that the in GU occurrence and recurrence of the process, PWP significantly inhibited NF-κB expression in the nucleus, has anti-inflammatory effects. 2.1.3 NF-κB protein expression in Western blot protein quantitative results of Western-blot test results: ⑴ 16d when the sham group NF-κB protein expression 0.50 ± 0.03, and the normal group (0.40 ± 0.03), no significant difference (P gt; 0.05); MWR group was 10.66 ± 1.92, significantly higher than the normal group and sham group (all P lt; 0.01); the OMLZ group and the the PWP group of NF-κB protein expression of 4.75 ± 0.93 and 4.50 ± 1.00, respectively compared with MWR group was significantly lower (P lt; 0.01). The ⑵ 92d the MR group NF-κB protein expression to 8.16 ± 0.72, significantly higher than the MWR sham group (0.61 ± 0.05) (P gt; 0.05); OMLZ group and the PWP group, respectively, to 4.26 ± 0.56 and 3.82 ± 0.42, and MR group, significantly lower (P lt; 0.01). Show that the in GU occurrence and recurrence process, activation of NF-κB protein expression increased, PWP can inhibit the expression of NF-κB protein. 2.2 PWP cellular localization of GU and its recurrence rat gastric tissue and plasma of IL-8 in 2.2.1 IL-8-positive cells expressing IL-8 positive cells are mainly located in the cytoplasm of mucosal epithelial cells stained brown or brown-black. Modeling 16d, the normal group and sham-operated rats, the gastric mucosa and submucosa expression twisted MWR group was significantly increased expression OMLZ group and the PWP group MWR group were reduced. 92d time, MWR, normal and sham group similar to the expression, are less MR group expression MWR group increased significantly, while OMLZ group and the PWP group was significantly lower than the MR group. ⑴ 16d 2.2.2 IL-8-positive cells express the results of immunohistochemistry, the normal group and sham group IL-8-positive cells express very little, 10.63 ± 0.04 and 10.91 ± 0.71, respectively, the two groups showed no significant difference (P gt; 0.05); MWR group to 41.73 ± 0.04, compared with normal group and sham surgery group significantly with an increase in (all P lt; 0.01); OMLZ group and the PWP group expression, respectively, 21.65 ± 1.96 and 22.01 ± 0.05, significantly with lower than MWR group (P lt; 0.01); the ⑵ 92d, MWR group and the normal group and sham group IL-8 expression of positive cells was 11.45 ± 0.79,12.15 ± 0.56 and 12.92 ± 0.72, respectively, between the three was no significant difference (both P gt; 0.05); compared with MWR MR group expressed positive cells increased significantly (P lt; 0.01), for 61.71 ± 1.43; OMLZ group and the PWP group were 38.21 ± 3.07 and 37.06 ± 1.82, compared with MR were significantly lower (all P lt; 0.01). Show that PWP can reduce the inflammatory cytokines IL-8 positive cells thereby reducing the GU recurrence. 2.2.3 IL-8 protein expression detection results of Western-blot Western blot protein quantification results show: ⑴ 16d when the normal group and sham group IL-8 protein expression were 10.45 ± 1.97 and 10.85 ± 1.23 and 39.63 ± MWR 4.27, significantly enhanced (P lt; 0.01); OMLZ group and the PWP group protein than the normal group was 18.33 ± 2.31 and 19.92 ± 3.30, respectively, compared with MWR group was significantly lower (all P lt; 0.01). ⑵ 92d, MWR group of IL-8 protein expression compared with normal and sham group without significantly with differences (all P gt; 0.05), the MR group IL-8 protein expression was 50.22 ± 3.05, compared to the MWR group (12.53 ± 3.50) significantly with enhanced ( P lt; 0.01); and OMLZ group and the PWP group were 32.23 ± 3.49 and 31.06 ± 3.28, significantly lower than the MR group (all P lt; 0.01). Show that the PWP by reducing IL-8 protein expression of anti-GU recurrence. 2.2.4 PWP recurrence GU rat plasma inflammatory cytokines IL-8 were radioimmunoassay, GU relapse MR plasma IL-8 content was 0.48 ± 0.07 ng / ml, compared with MWR group (0.14 ± 1.18 ng / ml) significantly increased (P lt; 0.01), OMLZ group and the PWP group, respectively, to 0.36 ± 0.43 ng / ml and 0.34 ± 0.03 ng / ml, compared to the MR group were significantly significantly reduce the (average P lt; 0.01) . Show that PWP to play anti GU recurrence role by lower plasma IL-8 content. 2.3 PWP GU and its recurrence rat gastric tissue and plasma TNF-α 2.3.1 TNF-α-positive cells in the expression of cell positioning modeling 16d, in the normal group and sham-operated rats, TNF-α positive cells in the gastric mucosa at the bottom of the cytoplasm and submucosa no or little expression. MWR group was significantly higher than the normal group and sham group OMLZ group and the PWP group expression MWR group to reduce. MWR group expression and expression sites and the expression level of the normal group and the sham group 92d when the MR group compared with the MWR group increased significantly in the the ulcer edge mucous membrane cytoplasmic expression, OMLZ group and the PWP group was significantly lower than the MR group. 2.3.2 TNF-α-positive cells expressed immunohistochemical results ⑴ 16d, the normal group and sham group only a small amount of TNF-α-positive cells expressed 3.90 ± 0.18 and 0.40 ± 0.04, respectively, the two groups showed no significant difference (P gt; 0.05), compared with the normal group, MWR group increase (21.59 ± 4.58) (P lt; 0.01), OMLZ group and the PWP group was 14.05 ± 2.55 and 13.67 ± 3.12, compared to the MWR group significantly with reduce ( all P lt; 0.01); the ⑵ 92d MWR group and the normal group and sham surgery group of TNF-α-positive cell number without significantly with sexual difference (all P gt; 0.05), the MR group-positive cell number to 40.13 ± 8.48, compared with MWR group (0.44 ± 0.05) was significantly increased (P lt; 0.01); OMLZ group and the PWP group were 19.92 ± 3.72 and 18.88 ± 4.17, were significantly lower than the MR group (all P lt; 0.01). That PWP by reducing the number of positive cells in the inflammatory cytokines TNF-α expression of anti-GU and its recurrence. 2.3.3 TNF-α protein expression in Western-blot test results of Western blot protein quantification results showed that: ⑴ 16d, the MWR of TNF-α protein expression to 19.80 ± 4.09, compared with normal and sham group increased significantly (all P lt; 0.01 ); OMLZ group and the PWP group were 13.02 ± 2.58 and 12.36 ± 2.49, compared to the MWR group significantly with reducing (both P lt; 0.01); 92d time, MWR group with normal and sham operation group protein expression without significantly with difference (P gt; 0.05); the MR group TNF-α protein expression to 34.64 ± 5.36, compared to the MWR group (0.38 ± 0.04) significantly with enhanced (P lt; 0.01); OMLZ group and the PWP group was 17.43 ± 2.35 and 16.96 ± 3.30 and MR group compared the difference was statistically significant (all P lt; 0.01). Show that the the PWP by reducing TNF-α protein expression of anti-GU and its recurrence role. 2.3.4 PWP inflammatory cytokines TNF-α levels in recurrence GU plasma radioimmunoassay results, GU relapse MR plasma TNF-α content of 23.74 ± 2.70 fmol / ml, compared to the MWR group (12.07 ± 1.73 fmol / ml) was significantly higher (P lt; 0.01), OMLZ group and the PWP group were 14.00 ± 2.02 fmol / ml and 13.55 ± 1.95 fmol / ml, compared to the MR group were significantly decreased (all P lt; 0.01). Showed that the PWP to play anti GU recurrence role by reducing the content of plasma TNF-α. Conclusion: the PWP able to effectively promote ulcer healing induced by acetic acid in rats with experimental GU, with anti GU recurrence role. 2. PWP anti GU recurrence role in their to play an anti-inflammatory effect by inhibiting NF-κB activation, reduced IL-8, TNF-α overexpression.
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