Microscopic appearance of the distal esophageal segment in all groups. A thickened wall and narrowed lumen in a rat with caustic esophageal injury from the burn group ( ► Fig. 2a ). In the burn þ 

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Context 1

... all the groups, NaOH and saline solutions were infused for 60 seconds, and the solutions were aspirated with a catheter. After removal of the sutures and withdrawal of the catheters, the layers of the abdominal wall were closed. Rats were fed ad libitum 6 hours after the procedure and were humanely killed on Day 22 after the burn. The distal esophageal segments were removed for biochemical and histologic analyses. The samples were sagittally divided into equal halves. The proximal half was kept frozen at – 80°C for biochemical analysis and the distal half was placed in a 10% formaldehyde solution for histologic analysis. Transverse sections of the esophageal specimens (4 – 5 μm) were obtained and were stained with hematoxylin and eosin stain and Masson trichrome stain. Histopathologic sections were examined by a single pathologist blinded to the groups. SI, which was de fi ned as the esophageal wall thickness divided by the lumen diameter, was calculated for each rat, and the mean SIs for all of the groups was compared. Histopathologic damage scores were based on collagen accumulation in the submucosal and muscular layers and injury to the muscularis mucosal and muscular layers 18 ( ► Table 1 ). MPO activity was measured in a manner similar to a method used by Kruidenier et al. 19 The tissue was homog- enized and then incubated in 0.5% hexadecyltrimethylam- monium bromide (pH 5.5); then, 0.026% ortho- dianisidinedihyrochloride and 0.018% H 2 O 2 was added the homogenate. The reaction kinetics at room temperature was followed for 30 minutes at a wavelength of 450 nm. Speci fi city for the reaction was con fi rmed by using sodium azide (0.1 mM). All analyses were conducted in at least two independent trials. MDA levels were analyzed spectrophotometrically. Brie fl y, 0.2 mL of each sample was mixed with 0.8 mL of tamponaded phosphate and 25 μL of butylated hydroxytoluene (pH 7.4). Trichloroacetic acid and hydroxybarbituric acid were added to the prepared solution. The mixture was kept on ice for 2 hours and then was centrifuged at 2,000 Â g for 15 minutes at 25°C. Then, a mixture of 0.075 mL ethylenediaminetetraacetic acid and 0.25 mL of 1% thiobarbituric acid was added to each mL of the prepared sample solution. The supernatant was boiled in water for 15 minutes and then cooled at room temperature. Spectrometric measurement was performed on the supernatant at a wavelength of 532 nm. The results were recorded in nmol/mg. NO levels were measured for each sample by using Griess reagent according to a method previously published by Moshage. 20 A nitrate reductase reaction was performed to convert nitrate to nitrite in the samples. Nitrogen compo- nents that reacted with the Griess reagent formed a purple color. Zinc sulfate was added to the samples; then, the samples were centrifuged at 10,000 Â g for 5 minutes. Measurements were performed by using an azo chromatographic spectrophotometer set at a wavelength of 450 nm. Values were recorded in mmol/mg. TNF- α and IL-6 levels were measured by an immunoenzy- matic ELISA technique (Quantikine ELISA Kit, R&D Systems, Minneapolis, Minnesota, United States). The minimum de- tectable values were 0.12 pg/mL for TNF- α and 0.03 pg/mL for IL-6. For comparisons of quantitative data between the three groups, ANalysis Of VAriance (ANOVA) was performed, cou- pled with post hoc Tukey tests. Data with nonnormal distri- butions were evaluated by using the Kruskal – Wallis test, and the Dunn test was employed as a post-hoc test. A p value < 0.05 was considered statistically signi fi cant. Two rats in the groups that received esophageal burns died within the fi rst 24 hours of the experiment as a result of aspirating the corrosive substance. These animals were ex- cluded from further analysis, and our results are thus based on the remaining 21 rats. Only, rats in the burn group showed signi fi cant weight loss. When the three groups of rats were compared for changes in body weight, signi fi cant differences between the groups were detected ( p < 0.05) ( ► Fig. 1 ). Upon macroscopic assessment, minimal adhesions were detected in the burn þ DMSO group, and the esophagi of the control group appeared normal without evidence of any adhesions. However, in the group of rats subjected to esophageal burns that did not receive DMSO, extremely sticky adhesions and hypertrophic esophageal tissue were detected. Macroscopic assessments were compatible with the histopathologic fi ndings ( ► Fig. 2 ). Histopathologic damage scores were signi fi cantly higher in the burn group compared with the burn þ DMSO and control groups, ( p < 0.05). The SI was signi fi cantly higher in the burn group relative to the other groups, ( p < 0.05). There were no statistically signi fi cant differences between the burn þ DMSO and control groups for the histopathologic damage scores or the SIs ( ► Fig. 3 ). In the burn group, MDA, NO ( ► Fig. 4 ), TNF- α , and IL-6 ( ► Fig. 5 ) levels were signi fi cantly higher relative to the other groups ( p < 0.05). However, no statistically signi fi cant differences were detected between the burn þ DMSO and control groups for any of these parameters. MPO levels were signi fi cantly increased in the burn and burn þ DMSO groups relative to the control group ( p < 0.05) and there was no signi fi cant difference in the MPO levels when comparing the burn þ DMSO and burn groups ( p 1⁄4 0.23) ( ► Fig. 6 ). The histopathologic and biochemical fi ndings for all the groups are shown in ► Table 2 . In our study, DMSO demonstrated a protective effect on stricture formation after caustic esophageal injury. DMSO signi fi cantly decreased the severity of the pathologic lesions and caused a marked reduction in the biochemical oxidative parameters. These effects could be associated with its antioxidant and anti-in fl ammatory activity. Corrosive esophageal burns represent a serious problem, especially in children. One of the main targets of esophageal burn therapy is to facilitate wound healing and to prevent potential stricture formation. 3,5,6,21 During the early phase after oral intake of a corrosive agent, in fl ammatory reactions occur, and free oxygen radicals are released. The formation of free oxygen radicals (i.e., NO and MDA) and cytokines (i.e., IL-6 and TNF- α ) during the acute phase plays a critical role in the development of tissue damage. Moreover, increases in tissue MPO activity indicate an accumulation of polymorphonuclear lymphocytes, which correlates with the extent of in fl ammation and damage. 22 The degree of damage also correlates with the severity of stricture formation. 23,24 Various treatment modalities have been identi fi ed that can reduce esophageal tissue damage and prevent stricture formation; however, a widely accepted medical treatment protocol has yet to be developed. Studies have been focused primarily on decreasing esophageal tissue damage and collagen formation, and various agents such as heparin, erythropoietin, ketotifen, vitamin E, ebselen, dexpanthenol, and ozone have been used for this purpose. 1,2,21,24 – 27 DMSO has been shown to ameliorate the acute phase symptoms in corrosive esophagitis. 12 However, our study investigated the effects of DMSO on esophageal stricture formation that is a late-stage complication. In our study, DMSO was administered during the fi rst week of the esophageal burn. At the end of the third week, DMSO signi fi cantly decreased the SI, the histopathologic damage score, and the levels of MDA, NO, IL-6, and TNF- α without having a signi fi cant impact on tissue MPO levels. DMSO has many different pharmacologic effects, including anti-in fl ammatory, antioxidant, cryoprotective, collagen-dissolving, and proteoglycan synthesis-inhibiting effects. On the basis of these properties, DMSO has been used successfully for the treatment of many diseases, via different routes, such as intravesically (interstitial cystitis), topically (cutaneous amyloidosis), orally (systemic amyloidosis), and parenterally (cerebral edema). 10 The fi rst 4 days after the induction of an esophageal burn are considered the acute necrotic phase. During this period, free oxygen radicals are released in response to an increase in cytokines, which results in tissue damage. The severity of tissue damage determines the degree of stricture formation. 23 Increases in the levels of cytokines IL-6 and TNF- α as a result of in fl ammation have been well characterized. In addition, the free-oxygen-radical-related MDA is accepted as a reliable indicator of lipid peroxidation and tissue damage associated with oxidative stress. 28 Tissue NO 29,30 and MPO activities have also been used as robust indicators of in fl ammation and tissue damage. 22 Suppression of the acute phase symptoms in patients with corrosive esophagitis using antioxidants and anti-in fl ammatory agents has consistently been shown to be an important treatment modality. DMSO is a hydrogen-bound disrupter, a cell-differentiat- ing agent and a hydroxyl radical scavenger. Antioxidant and anti-in fl ammatory properties of DMSO have been demonstrated in previous studies. 13 – 15 Cytokines enable the induction and regulation of immunologic and in fl ammatory responses. In our study, DMSO signi fi cantly decreased the levels of IL-6 and TNF- α . These fi ndings are consistent with results reported by Essani et al 8 and Kloesch et al. 9 Because of its antioxidant and anti-in fl ammatory effects, DMSO ...

Context 2

... all the groups, NaOH and saline solutions were infused for 60 seconds, and the solutions were aspirated with a catheter. After removal of the sutures and withdrawal of the catheters, the layers of the abdominal wall were closed. Rats were fed ad libitum 6 hours after the procedure and were humanely killed on Day 22 after the burn. The distal esophageal segments were removed for biochemical and histologic analyses. The samples were sagittally divided into equal halves. The proximal half was kept frozen at – 80°C for biochemical analysis and the distal half was placed in a 10% formaldehyde solution for histologic analysis. Transverse sections of the esophageal specimens (4 – 5 μm) were obtained and were stained with hematoxylin and eosin stain and Masson trichrome stain. Histopathologic sections were examined by a single pathologist blinded to the groups. SI, which was de fi ned as the esophageal wall thickness divided by the lumen diameter, was calculated for each rat, and the mean SIs for all of the groups was compared. Histopathologic damage scores were based on collagen accumulation in the submucosal and muscular layers and injury to the muscularis mucosal and muscular layers 18 ( ► Table 1 ). MPO activity was measured in a manner similar to a method used by Kruidenier et al. 19 The tissue was homog- enized and then incubated in 0.5% hexadecyltrimethylam- monium bromide (pH 5.5); then, 0.026% ortho- dianisidinedihyrochloride and 0.018% H 2 O 2 was added the homogenate. The reaction kinetics at room temperature was followed for 30 minutes at a wavelength of 450 nm. Speci fi city for the reaction was con fi rmed by using sodium azide (0.1 mM). All analyses were conducted in at least two independent trials. MDA levels were analyzed spectrophotometrically. Brie fl y, 0.2 mL of each sample was mixed with 0.8 mL of tamponaded phosphate and 25 μL of butylated hydroxytoluene (pH 7.4). Trichloroacetic acid and hydroxybarbituric acid were added to the prepared solution. The mixture was kept on ice for 2 hours and then was centrifuged at 2,000 Â g for 15 minutes at 25°C. Then, a mixture of 0.075 mL ethylenediaminetetraacetic acid and 0.25 mL of 1% thiobarbituric acid was added to each mL of the prepared sample solution. The supernatant was boiled in water for 15 minutes and then cooled at room temperature. Spectrometric measurement was performed on the supernatant at a wavelength of 532 nm. The results were recorded in nmol/mg. NO levels were measured for each sample by using Griess reagent according to a method previously published by Moshage. 20 A nitrate reductase reaction was performed to convert nitrate to nitrite in the samples. Nitrogen compo- nents that reacted with the Griess reagent formed a purple color. Zinc sulfate was added to the samples; then, the samples were centrifuged at 10,000 Â g for 5 minutes. Measurements were performed by using an azo chromatographic spectrophotometer set at a wavelength of 450 nm. Values were recorded in mmol/mg. TNF- α and IL-6 levels were measured by an immunoenzy- matic ELISA technique (Quantikine ELISA Kit, R&D Systems, Minneapolis, Minnesota, United States). The minimum de- tectable values were 0.12 pg/mL for TNF- α and 0.03 pg/mL for IL-6. For comparisons of quantitative data between the three groups, ANalysis Of VAriance (ANOVA) was performed, cou- pled with post hoc Tukey tests. Data with nonnormal distri- butions were evaluated by using the Kruskal – Wallis test, and the Dunn test was employed as a post-hoc test. A p value < 0.05 was considered statistically signi fi cant. Two rats in the groups that received esophageal burns died within the fi rst 24 hours of the experiment as a result of aspirating the corrosive substance. These animals were ex- cluded from further analysis, and our results are thus based on the remaining 21 rats. Only, rats in the burn group showed signi fi cant weight loss. When the three groups of rats were compared for changes in body weight, signi fi cant differences between the groups were detected ( p < 0.05) ( ► Fig. 1 ). Upon macroscopic assessment, minimal adhesions were detected in the burn þ DMSO group, and the esophagi of the control group appeared normal without evidence of any adhesions. However, in the group of rats subjected to esophageal burns that did not receive DMSO, extremely sticky adhesions and hypertrophic esophageal tissue were detected. Macroscopic assessments were compatible with the histopathologic fi ndings ( ► Fig. 2 ). Histopathologic damage scores were signi fi cantly higher in the burn group compared with the burn þ DMSO and control groups, ( p < 0.05). The SI was signi fi cantly higher in the burn group relative to the other groups, ( p < 0.05). There were no statistically signi fi cant differences between the burn þ DMSO and control groups for the histopathologic damage scores or the SIs ( ► Fig. 3 ). In the burn group, MDA, NO ( ► Fig. 4 ), TNF- α , and IL-6 ( ► Fig. 5 ) levels were signi fi cantly higher relative to the other groups ( p < 0.05). However, no statistically signi fi cant differences were detected between the burn þ DMSO and control groups for any of these parameters. MPO levels were signi fi cantly increased in the burn and burn þ DMSO groups relative to the control group ( p < 0.05) and there was no signi fi cant difference in the MPO levels when comparing the burn þ DMSO and burn groups ( p 1⁄4 0.23) ( ► Fig. 6 ). The histopathologic and biochemical fi ndings for all the groups are shown in ► Table 2 . In our study, DMSO demonstrated a protective effect on stricture formation after caustic esophageal injury. DMSO signi fi cantly decreased the severity of the pathologic lesions and caused a marked reduction in the biochemical oxidative parameters. These effects could be associated with its antioxidant and anti-in fl ammatory activity. Corrosive esophageal burns represent a serious problem, especially in children. One of the main targets of esophageal burn therapy is to facilitate wound healing and to prevent potential stricture formation. 3,5,6,21 During the early phase after oral intake of a corrosive agent, in fl ammatory reactions occur, and free oxygen radicals are released. The formation of free oxygen radicals (i.e., NO and MDA) and cytokines (i.e., IL-6 and TNF- α ) during the acute phase plays a critical role in the development of tissue damage. Moreover, increases in tissue MPO activity indicate an accumulation of polymorphonuclear lymphocytes, which correlates with the extent of in fl ammation and damage. 22 The degree of damage also correlates with the severity of stricture formation. 23,24 Various treatment modalities have been identi fi ed that can reduce esophageal tissue damage and prevent stricture formation; however, a widely accepted medical treatment protocol has yet to be developed. Studies have been focused primarily on decreasing esophageal tissue damage and collagen formation, and various agents such as heparin, erythropoietin, ketotifen, vitamin E, ebselen, dexpanthenol, and ozone have been used for this purpose. 1,2,21,24 – 27 DMSO has been shown to ameliorate the acute phase symptoms in corrosive esophagitis. 12 However, our study investigated the effects of DMSO on esophageal stricture formation that is a late-stage complication. In our study, DMSO was administered during the fi rst week of the esophageal burn. At the end of the third week, DMSO signi fi cantly decreased the SI, the histopathologic damage score, and the levels of MDA, NO, IL-6, and TNF- α without having a signi fi cant impact on tissue MPO levels. DMSO has many different pharmacologic effects, including anti-in fl ammatory, antioxidant, cryoprotective, collagen-dissolving, and proteoglycan synthesis-inhibiting effects. On the basis of these properties, DMSO has been used successfully for the treatment of many diseases, via different routes, such as intravesically (interstitial cystitis), topically (cutaneous amyloidosis), orally (systemic amyloidosis), and parenterally (cerebral edema). 10 The fi rst 4 days after the induction of an esophageal burn are considered the acute necrotic phase. During this period, free oxygen radicals are released in response to an increase in cytokines, which results in tissue damage. The severity of tissue damage determines the degree of stricture formation. 23 Increases in the levels of cytokines IL-6 and TNF- α as a result of in fl ammation have been well characterized. In addition, the free-oxygen-radical-related MDA is accepted as a reliable indicator of lipid peroxidation and tissue damage associated with oxidative stress. 28 Tissue NO 29,30 and MPO activities have also been used as robust indicators of in fl ammation and tissue damage. 22 Suppression of the acute phase symptoms in patients with corrosive esophagitis using antioxidants and anti-in fl ammatory agents has consistently been shown to be an important treatment modality. DMSO is a hydrogen-bound disrupter, a cell-differentiat- ing agent and a hydroxyl radical scavenger. Antioxidant and anti-in fl ammatory properties of DMSO have been demonstrated in previous studies. 13 – 15 Cytokines enable the induction and regulation of immunologic and in fl ammatory responses. In our study, DMSO signi fi cantly decreased the levels of IL-6 and TNF- α . These fi ndings are consistent with results reported by Essani et al 8 and Kloesch et al. 9 Because of its antioxidant and anti-in fl ammatory effects, DMSO ...