Dexmedetomidine protects rats from postoperative cognitive dysfunction via regulating the GABAB R-mediated cAMP-PKA-CREB signaling pathway.

This work attempts to discuss whether dexmedetomidine (Dex) can protect rats from postoperative cognitive dysfunction (POCD) through regulating the γ-aminobutyric acid-B receptor (GABAB R)-mediated cyclic adenosine monophosphate (cAMP) - protein kinase A (PKA) - cAMP-response element binding (cAMP-PKA-CREB) signaling pathway. Sprague-Dawley rats were divided into a non-surgical group (Control), a surgical group (Model), a surgical group treated with Dex (Model + Dex), a surgical group treated with GABAB R antagonist (Model + CGP 35348) and a surgical group treated with Dex and GABAB R agonist (Model + Dex + Baclofen). Cognitive and memory functions were evaluated by Y-maze test and open-field test. The neuronal morphology of the hippocampus was observed by hematoxylin and eosin staining and neuronal apoptosis was by terminal deoxynucleotidyl transferase-mediated dUTP-biotin nick-end labeling method. Inflammatory factors and cAMP levels were detected by enzyme-linked immunosorbent assay while expressions of GABAB R and PKA-CREB pathway-related molecules by Western blot. Compared with control rats, the model rats exhibited reduced alternation rates with a prolonged time spent in the central zone; meanwhile, levels of tumor necrosis factor-α and interleukin-1ß and the apoptotic index, as well as GABAB R1 and GABAB R2 expressions were increased in the model rats, but the cAMP-PKA-CREB pathway was inhibited (all P < 0.05). When treated with either Dex or CGP 35348, the surgical rats displayed an opposite tendency concerning the above factors as compared to the model rats (all P < 0.05). Furthermore, Baclofen, the agonist of GABAB R, could reverse the protective effect of Dex against POCD in rats. Dex protects rats from POCD possibly via suppressing GABAB R to up-regulate the cAMP-PKA-CREB signaling pathway, thereby alleviating the hippocampal inflammation caused by surgical trauma.

Effects of maternal enflurane exposure on NR2B expression in the hippocampus of their offspring

This work aims to study the pathogenesis of learning and memory impairment in offspring rats resulting from maternal enflurane anesthesia by focusing on the expression of the N-methyl-d-aspartic acid receptor subunit 2B (NR2B) in the hippocampus of the offspring. Thirty female Sprague-Dawley rats were randomly divided into three groups: control (C group), 4 h enflurane exposure (E1 group), and 8 h enflurane exposure (E2 group) groups. Eight to ten days after the initiation of pregnancy, rats from the E1 and E2 groups were allowed to inhale 1.7% enflurane in 2 L/min oxygen for 4 h and 8 h, respectively. Rats from the C group were allowed to inhale 2 L/min of oxygen only. The Morris water maze was used to assay the learning and memory function of the offspring on postnatal days 20 and 30. RT-PCR and immunohistochemistry assays were then used to measure the mRNA levels and protein expression of NR2B, respectively. Relative to offspring rats from the C group, those from the E1 and E2 groups exhibited longer escape latencies, lesser number of crossings over the platform, and less time spent in the target quadrant in the spatial exploration test (P < 0.05). In addition, the mRNA and protein expression levels of NR2B in the hippocampus of offspring rats in the E1 and E2 groups were down-regulated (P < 0.05). No significant differences between the E1 and E2 groups were observed (P > 0.05) in terms of mRNA levels and protein expression of NR2B. The cognitive function of the offspring is impaired when maternal rats are exposed to enflurane during early pregnancy. A possible mechanism of this effect is related to the down-regulation of NR2B expression.

Este trabalho objetiva o estudo da patogênese de deficiência no aprendizado e memória de prole de ratos resultante da anestesia maternal por enflurano, por meio da expressão da subunidade 2B do receptor do ácidoN-metil-D-aspártico (NR2B) no hipocampo dos filhotes. Dividiram-se, aleatoriamente, 30 fêmeas de ratos Sprague-Dawley em três grupos: controle (grupo C), exposição ao enflurano por 4 h (grupo E1) e por 8 h (grupo E2). De oito a 10 dias após o início da gravidez, os ratos dos grupos E1 e E2 inalaram enflurano 1,7% em 2 L/min de oxigênio, por 4 h e 8 h, respectivamente. Ratos do grupo C inalaram apenas 2 L/min de oxigênio. O labirinto de água de Morris foi empregado para analisar as funções de aprendizado e memória da cria em 20 e 30 dias após o nascimento. Utilizaram-se ensaios de RT-PCR e de imuno-histoquímica para medir os níveis de mRNA e expressão da proteína do NR2B, respectivamente. Em comparação com os ratos controle do grupo C, aqueles dos grupos E1 e E2 exibiram latências de escape mais longas, menor número de travessias na plataforma e menos tempo gasto no quadrante alvo no teste de exploração espacial (P < 0,05). Adicionalmente, os níveis de expressão de mRNA e de proteína do NR2B no hipocampo dos filhotes nos grupos E1 e E2 estavam reduzidos (P < 0,05). Não se observaram diferenças significativas entre os grupos E1 e E2 (P < 0,05) quanto aos níveis de mRNA e à expressão de proteína de NR2B. A função cognitiva dos filhotes é prejudicada quando as mães são expostas ao enflurano durante o início da gravidez. O mecanismo possível para esse efeito está relacionado à diminuição na expressão de NR2B.

Regulation on the expression of Clara cell secretory protein in the lungs of the rats with acute lung injury by growth hormone.

BACKGROUND: Clara cell secretory protein (CC16) is an important lung derived protective factor and may play an important role on the pathogenesis of acute lung injury (ALI) induced by endotoxemia. Growth hormone (GH) is an important anabolism hormone secreted by GH cells of the hypophysis. Previous research showed that GH would significantly exacerbate ALI induced by endotoxemia, but the mechanism is not very clear yet. Whether the effects are related to CC16 or not is undetermined. METHODS: One hundred and twelve male Sprague-Dawley rats were randomly divided into an ALI group and a GH group. The rats in the two groups were subdivided into seven subgroups, according to injection with lipopolysaccharides (LPS) or not, then according to different intervals of time after LPS injection; 0 hour (pre-injection of LPS, acted as control group), 0.5 hour, 1 hour, 2 hours, 4 hours, 6 hours and 24 hours for subgroups. Pulmonary alveolar septa area density (PASAD) and ploymorphonuclear cells (PMN) in the lungs were analyzed morphometrically. The levels of tumor necrosis factor (TNF) and interleukin 6 (IL-6) were determined by radioimmunoassay. To analyze the expression and activation of nuclear factor kappa B (NF-κB), the numbers of NF-κB positive cells in lungs were counted after immunofluorescence staining, and the levels of NF-κB inhibitory protein-α (IκB-α) in lung homogenates of rats were detected by Western blotting. The expression levels of CC16 mRNA in lungs of the rats with ALI were determined by semi-quantitative reverse transcription polymerase chain reaction (RT-PCR). The levels of CC16 protein in lung homogenates were detected by Western blotting. RESULTS: Half an hour after LPS injury both the PASAD and PMN numbers in lungs of the rats with ALI began to increase significantly and peaked at 6-hour post-injury. They then began to recover and reached normal levels at 24-hour post-injury. Both the PASAD and PMN numbers in the GH group increased more significantly than those in the ALI group. The levels of TNF in lungs of the rats with ALI homogenates increased significantly 0.5-hour post-injury, peaked at 1-hour and maintained a high level until 6 hours then gradually recovered. The content of TNF in the GH group lung homogenates increased more significantly than in the ALI group post-injury. The contents of IL-6 in rat lung homogenates began to increase significantly at 1-hour post-injury, peaked at 4 hours then gradually returned to normal levels by 6 hours post-injury. The levels of IL-6 in the lung homogenates of the GH group were higher than in the ALI group at different time intervals post-injury. The number of NF-κB positive cells increased dramatically at 0.5-hour post-injury, and the fluorescence intensity was enhanced. Both peaked at 4-hour post-injury. The number of NF-κB positive cells and the enhanced intensity of fluorescence began to decrease from 6-hour post-injury, but the number of NF-κB cells at 24 hours post-injury was still higher than in the control group. The number of NF-κB cells in lungs in the GH group was significantly higher than in the LPS group at the different time intervals post-injury. The IκB-α expression in lungs of the rats with ALI homogenates decreased dramatically 0.5-hour post-injury, reaching a nadir at 4-hour post-injury and then began to recover. The levels of IκB-α in GH group were significantly lower than those in ALI group. Both the levels of CC16 mRNA and protein in lungs of the rats with ALI began to decrease significantly 0.5-hour post-injury, reached a nadir at 6 hours, and then began to recover. Both the expression of CC16 mRNA and CC16 protein in the GH group were significantly lower than those in the ALI group at the different time intervals post-injury. Correlation analysis indicates that CC16 correlates significantly with all the indices mentioned above. CONCLUSIONS: Down-regulation of CC16 expression plays a critical role in the pathogenesis of acute lung injury induced by endotoxemia. The application of GH can exacerbate the lung injury induced by endotoxemia through down-regulating the expression of CC16.

[The effect of growth hormone on Clara cell in acute lung injury of rats].

OBJECTIVE: To study the effect of growth hormone (Gh) on Clara cell in acute lung injury (ALI) rats during stress responsive stage. METHODS: One hundred and twelve male Sprague-Dawley (SD) rats were randomly divided into ALI group and recombinant human Gh (rhGh) group. The rats in two groups were subdivided into seven subgroups of different time intervals after lipopolysaccharide (LPS) injection:0 (pre-injection of LPS, acted as control group), 0.5, 1, 2, 4, 6 and 24 hours subgroups respectively. Immunohistochemistry and electron microscope were used to investigate the number and morphological changes in Clara cells in ALI rats. RESULTS: Half of an hour after LPS injection, ALI was induced. The extent of lung injury increased with the time increased after LPS injection,peaked at 6 hours, then the extent of lung injury began to recess. The dynamic changes in lung injury of rats in rhGh group were similar to those in ALI group, but the extent of injury was more severe than that in ALI group at different time intervals. The Clara cell counts decreased significantly 1 hour after LPS injection, reaching the lowest at 6 hours, then began to recover 24 hours after LPS injection with statistically significant difference (all P<0.01). The counts of Clara cells in rhGh group decreased more significantly than that in ALI group at different time intervals after LPS injection, especially at 24 hours.The ultrastructure of Clara cell showed significant damage 24 hours after LPS injection. Compared with ALI group,the ultrastructure of Clara cell in rhGh group were more damaged 24 hours after LPS injection. CONCLUSION: rhGh could deteriorate the damage of Clara cell in acute lung injury rats induced by endotoxemia during stress response stage.

[Effect of growth hormone on acute lung injury].

OBJECTIVE: To study the effect of growth hormone (GH) on acute lung injury (ALI) induced by endotoxemia, and its potential therapeutic mechanism. METHODS: One hundred and twelve male Sprague-Dawley rats were randomly divided into ALI group and GH group. The rats in two groups were further divided into seven subgroups determined by the length of interval after lipopolysaccharide (LPS) challenge: 0 (before injection of LPS, served as control group), 0.5, 1, 2, 4, 6 and 24 hours subgroups respectively. Pulmonary alveolar septum area density (PASAD) and the number of neutrophil in lungs of rats were analyzed morphometrically. The levels of tumor necrosis factor (TNF) and interleukin-6 (IL-6) were determined by radioimmunoassay. The expression and activation of nuclear factor-kappaB (NF-kappaB) were analyzed, NF-kappaB positive cells in lungs were counted after immunofluorescence staining, and the levels of NF-kappaB inhibition (I-kappaB alpha) in lung homogenates of rats were assayed by Western blot. RESULTS: Half an hour after intravenous injection of LPS, both the PASAD and the number of neutrophil in lungs of ALI rats began to increase, and peaked at 6 hours post-injury, then began to recover and reached the normal levels at 24 hours. The content of TNF in lung homogenates showed immediate elevation after LPS injection, becoming higher than that of the control after 0.5 hour, reaching the peak value at 1 hour, maintaining high levels until 6 hours, then gradually recovered. The content of TNF in lung homogenates of the GH group increased significantly more than that in the LPS group. The contents of IL-6 in rats' lung homogenates began to increase significantly 1 hour post-injury, peaked at 4 hours, then gradually returned to normal level 6 hours post-injury. The content of IL-6 in the lung homogenates of GH group was higher than that in the LPS group at different time intervals post-injury, showing significant difference at 0.5, 6 and 24 hours (P<0.05 or P<0.01). The number of NF-kappaB positive cells increased dramatically at 0.5 hour post-injury. The intensity of fluorescence was enhanced. Both of them peaked at 4 hours post-injury. The number of NF-kappaB positive cells and the enhanced intensity of fluorescence began to decrease at 6 hours post-injury. But the number of NF-kappaB cells at 24 hours post-injury was still larger than that in the control group. The number of NF-kappaB cells in lungs of GH group was significantly larger than that in the LPS group at different time intervals post-injury (P<0.05 or P<0.01). I-kappaB alpha contents in lung homogenates of ALI rats decreased dramatically 0.5 hour post-injury, nadir at 4 hours, and then began to recover. Correlation analysis indicated that PASAD, the levels of TNF and IL-6 and the extent of neutrophil infiltration in lung were positively correlated to the extent of the expression and the activation of NF-kappaB. CONCLUSION: The application of GH during early stage of endotoxin challenge can deteriorate the lung injury induced by LPS through enhancing the expression and activation of NF-kappaB, thus enhances the inflammatory response in lungs.