Lipopolysaccharide mediates immuno-pathological alterations in young chicken liver through TLR4 signaling.

BACKGROUND: Lipopolysaccharide (LPS) induces acute liver injury and the complex mechanisms include the activation of toll like receptor 4 (TLR4) signaling pathway in many species. However, immuno-pathological changes during TLR4 signaling under LPS stress in acute liver injury is poorly understood in avian species. The present investigation was therefore carried out to evaluate these alterations in TLR4 signaling pathway during acute liver injury in young chickens. RESULTS: After intraperitoneal injection of LPS or saline, liver samples were harvested at 0, 2, 6, 12, 24, 36, 72 and 120 h (n = 6 at each time point) and the microstructures were analyzed by hematoxylin and eosin (H&E) staining. Alanine aminotransferase (ALT) and caspase-3 enzyme activity was assessed by enzyme-linked immunosorbent assay (ELISA). Proliferative cell nuclear antigen (PCNA), single stranded DNA (ssDNA) and TLR4 protein expressions were determined by immunohistochemistry. Gene expressions of PCNA, caspase-3, caspase-8, TLR4 and its downstream molecules were analyzed by quantitative polymerase chain reaction (qPCR). LPS injection induced significantly higher ALT activity, severe fatty degeneration, necrotic symptoms, ballooning degeneration, congestion, enhanced inflammatory cell infiltration in liver sinusoids, decreased proliferation, increased apoptosis and significant up-regulation in TLR4 and its downstream molecules (MyD88, NF-κB, TNF-α, IL-1ß and TGF-ß) expression at different time points. CONCLUSIONS: This study indicated that TLR4 signaling and its downstream molecules along with certain cytokines play a key role in acute liver injury in young chickens. Hence, our findings provided novel information about the histopathological, proliferative and apoptotic alterations along with changes in ALT and caspase-3 activities associated with acute liver injury induced by Salmonella LPS in avian species.

Effects of lipopolysaccharide on the histomorphology and expression of toll-like receptor 4 in the chicken trachea and lung.

Endotoxin or lipopolysaccharide (LPS) exposure can cause injury to the respiratory airways and in response, the respiratory epithelia express toll-like receptors (TLRs) in many species. However, its role in the innate immunity in the avian respiratory system is poorly understood. The aim of the present study was to evaluate the effects of LPS on the chicken trachea and lung. After intraperitoneal LPS or saline injection, the trachea and lungs were harvested at 0, 12, 36 and 72 h (n = 6 at each time point) and histopathologically analysed using haematoxylin and eosin and periodic acid-Schiff staining, while TLR4 expression was determined by immunohistochemistry and secretory Immunoglobulin A (SIgA) levels by enzyme-linked immunosorbent assay. After LPS stimulation, we observed a remarkable decrease in the number of goblet cells along with obvious disruption and desquamation of the ciliated epithelium in the trachea, blurring of the boundary between pulmonary lobules, narrowed or indistinguishable lumen of the pulmonary atria and leukostasis in the lungs. Following LPS stimulation, TLR4 protein expression was up-regulated in both the trachea and the lungs and was found on the ciliated columnar cells as well as in the submucosa of the trachea, and in the lungs on parenchymal and immune cells. However, SIgA levels were only up-regulated in the trachea at 12 h following LPS stimulation. Hence, this report provides novel information about the effects of LPS on the microstructure of the lower respiratory tract and it is concluded that its intra-peritoneal administration leads to TLR4-mediated destruction of the tracheal epithelium and pulmonary inflammation along with increased SIgA expression in the tracheal mucosa.

The effect of visfatin on inflammatory reaction in uterus of LPS-induced rats / Efecto de visfatina en la reacción inflamatoria en el útero de ratas inducidas por LPS

The present study was to investigate the effects of visfatin on the morphological structure and function of the rat uterus during inflammation. The expression and distribution of visfatin, morphological structure, eosinophils (EOS), myeloperoxidase (MPO) and cytokines in the uterus of the LPS-induced rat were studied using hematoxylin-eosin staining (HE), immunohistochemical methods, western blots and enzyme-linked immunosorbent assay (ELISA). The present study showed that visfatin positive cells dispersed widely in the uterus, and strong positive staining was observed mainly in the cell cytoplasm. Compared with saline group, in visfatin group, more uterine glands were found, EOS increased, and the difference was significant (P<0.05), MPO reduced, and the difference was significant (P<0.01). In addition, visfatin was able to increase the secretion of IL-1b, IL-6, and TNF-a (P<0.01). Compared with LPS group, in vifatin+LPS group, the uterine glands of the lamina propria increased, the myometrium became thinner, the number of EOS and MPO reduced obviously, but the difference was not significant (P>0.05), and after LPS stimulated body, visfatin decrease the level of IL-1b, IL-6, TNF-a (P<0.01). The above results suggest that visfatin could affect the morphological structure of rat uterus; Visfatin could modulate the inflammatory response in rats' uterus by regulating the quantity of inflammatory cells, such as EOS and MPO, and the level of inflammatory cytokines, such as IL-1b, IL-6, TNF-a.

El objetivo del presente estudio fue investigar los efectos de la visfatina sobre la estructura morfológica y la función del útero de la rata durante la inflamación. Se estudiaron la expresión y distribución de la visfatina, la estructura morfológica, eosinófilos, mieloperoxidasa y citoquinas en el útero de rata mediante la tinción de H&E, métodos inmunohistoquímicos, Western blots y ELISA. El estudio mostró que las células visfatina positivas se dispersan ampliamente en el útero, junto a una fuerte tinción positiva, principalmente en el citoplasma de la célula. En comparación con el grupo control, en el grupo visfatina, se encontraron más glándulas uterinas, se observó un aumento de EOS y la diferencia fue significativa (p<0,05), MPO reducida siendo esta diferencia también significativa (p<0,01). Además, la visfatina fue capaz de aumentar la secreción de IL-1b, IL-6 y TNF-a (P<0,01). En comparación con el grupo LPS, visfatina+grupo LPS, las glándulas uterinas de la lámina propia aumentaron, se observó un miometrio más delgado, y número reducido de EOS y MPO, sin embargo, la diferencia no fue significativa (P>0,05). Después de estímulo LPS en el cuerpo, se registró un nivel menor de visfatina en IL-1b, IL-6, TNF-a (P<0,01). Los resultados anteriores sugieren que visfatina podría afectar a la estructura morfológica del útero de rata. Además, podría modular la respuesta inflamatoria en el útero mediante la regulación de la cantidad de células inflamatorias, tales como EOS y MPO.

The role of visfatin on the regulation of inflammation and apoptosis in the spleen of LPS-treated rats.

The purpose of the present study is to determine if visfatin is involved in inflammation or apoptosis induced by LPS in rat. Forty Wistar rats were divided into four groups: saline group, LPS group, visfatin group and Visfatin + LPS co-stimulated group. Spleen samples from each group of rats were collected for study. The spleen structure was examined by histological imaging. Apoptosis was evaluated with TUNEL reaction. Caspase-3 was detected with immunohistochemistry and western blot. The apoptosis-related genes were detected by qPCR and inflammatory cytokines were tested by ELISA. Our main findings were as follows. (1) Macrophages were markedly increased in the visfatin group compared with the saline group. This finding was confirmed when spleen samples were examined with western blot using CD68 antibody. (2) Visfatin promoted the expression of CD68 and caspase-3 in rat spleen, whereas visfatin could inhibit the expression of CD68 and activated caspase-3 in spleen of LPS-induced acute inflammation. (3) Visfatin had a pro-apoptotic effect on normal rat spleen, whereas it exerted an anti-apoptotic effect during LPS-induced lymphocytes apoptosis in rat spleen. Moreover, the effect of visfatin on cell apoptosis was mediated by the mitochondrial pathway. (4) Visfatin could modulate both the anti-inflammatory cytokines and pro-inflammatory cytokines in rat spleen, such as IL-10, IL-4, IL-6, TNF-α and IL-1ß. Taken together, we demonstrate that visfatin could participate in the inflammatory process in rat spleen by modulating the macrophages and inflammatory cytokines. Also, visfatin plays a dual role in the apoptosis in rat spleen, which is mediated by the mitochondrial pathway.

Distribution and expression of visfatin-positive cells in the spleen of lipopolysaccharide-stimulated piglets / Distribución y expresión de células visfatin positivas en el bazo de lechones estimulados con lipopolisacáridos

The histological changes in the spleen and the immunohistochemical expression of visfatin in lipopolysaccharide-stimulated piglets are reported to examine the relation between visfatin and inflammation. The results are as follows: (1) After LPS treated, the spleen displayed thicker capsules and trabecula, the thinner periarterial lymphatic sheath, and the more expandable splenic sinusoid, with an increase in the number of splenic nodules, lymphocytes, ellipsoids of the marginal zone, red blood cells and macrophagocytes. (2) Visfatin-positive cells were mainly distributed in the red pulp of the spleen, with less in splenic nodules and periarterial lymphatic sheath. In the LPS-treated group, the signal intensity and quantity of the visfatin-positive cells were significantly higher in the red pulp and the ellipsoids of the spleen (P<0.01), whereas lower in the periarterial lymphatic sheath. These results indicate that LPS stimulation induces inflammation, causing the histological changes of the piglet spleen and activating humoral immune response. Moreover, variation of visfatin in the spleen suggests that lymphocytes and macrophages are the potent source of visfatin which participates in the humoral immune response in the inflammation.

Se presentan los cambios histológicos en el bazo y la expresión inmunohistoquímica de visfatin en lechones estimulados mediante lipopolisacáridos (LPS) con el objetivo de estudiar la relación entre visfatin e inflamación. Los resultados fueron los siguientes: (1) Después del tratamiento por LPS se observaron en el bazo cápsulas más gruesas y trabéculas, una vaina linfática periarterial más delgada, y más sinusoides esplénicos expandible, con un aumento en el número de nódulos esplénicos, linfocitos, elipsoides de la zona marginal, como también un aumento de las células rojas de la sangre y los macrofagocitos. (2) Las células visfatina-positivas se distribuyeron principalmente en la pulpa roja del bazo, con una cantidad menor en los nódulos esplénicos y la vaina linfática periarterial. En el grupo tratado con LPS, la intensidad de la señal y número de células positivas fueron significativamente mayor en la pulpa roja y los elipsoides del bazo (P<0,01), mientras que estas fueron menores en la vaina linfática periarterial. Estos resultados indican que la estimulación con LPS induce la inflamación provocando cambios histológicos del bazo de los lechones y la activación de la respuesta inmune humoral. Por otra parte, la variación de visfatin en el bazo sugiere que los linfocitos y los macrófagos son una fuente potente de visfatin en la respuesta inmune humoral de la inflamación.

TLR4 is constitutively expressed in chick thymic epithelial cells.

Toll-like receptor 4 (TLR4) has been suggested to play a regulatory role in immune cell development; however, studies regarding the role of TLR4 in the development of the chick thymus are scarce. In this study, we investigated the distribution and expression pattern of TLR4 in normal chick thymi at different stages of development, in order to better understand the role of TLR4 in chick thymus development. We studied the thymi from 15 chicks, collected at days 7, 21 and 35 of age. The relative change in TLR4 mRNA expression in the chick thymus at different ages was determined by quantitative real-time PCR, and changes in protein expression were analyzed by immunohistochemistry and Western blotting. Furthermore, the distribution of TLR4 in the chick thymus was analyzed by immunohistochemistry, and compared with the distribution of TLR4 expression in juvenile female pigs (gilts). Our results indicated that TLR4 was constitutively expressed in the chick thymus. TLR4 was primarily expressed in the thymic cortico-medullary junction and the medulla, particularly in the epithelial cells of Hassall's corpuscles. The mRNA and protein expression level of TLR4 increased in the thymus with increasing age (p<0.05). Taken together, these results indicate that TLR4 is constitutively expressed by epithelial cells in the chick thymus, suggesting it may participate in thymic development by inducing factors affecting its development.

Distribution and morphology of argyrophilic cells in the digestive tract of the African ostrich.

We used light microscopy to elucidate the morphological features of argyrophilic cells in the digestive tract of the African ostrich (Struthio camelus). The results indicated that argyrophilic cells were found to be distributed among the epithelial cells of the mucosa or glands throughout the digestive tract, except for the esophagus; two types of argyrophilic cells were found; i.e., closed-type cells and cells with triangular or elongated shapes and with their apical cytoplasmic process in contact with the lumen (open-type cells); the greatest number of argyrophilic cells was found in the proventriculus, and the argyrophilic cell density gradually decreased from the proventriculus to the rectum; Furthermore, the number of argyrophilic cells in the duodenum and ileum was higher than that in the jejunum. This text still combined the characteristics that the argyrophilic cells in digestive tract of ostrich maybe related to different digestive function of different region and the basis of their morphology to carry on a discussion. It was speculated that argyrophilic cells in the digestive tract may have both endocrine and exocrine functions.