ABSTRACT
ObjectiveTo explore the effect and mechanism of the classic famous prescription Anmeidan （AMD） developed in the Qing Dynasty in regulating the hepatic neurotransmitters and circadian rhythm in the rat model of insomnia via the orexin-1 receptor （OX1R）/phosphatidylinositol-specific phospholipase Cβ-1 （PLCβ-1）/protein kinase Cα （PKCα）/extracellular signal-regulated kinase 1/2 （ERK1/2） signaling pathway. MethodSixty SPF-grade SD rats were randomized into blank， model， suvorexant （30 mg·kg-1·d-1）， and low-， medium-， and high-dose （4.55， 9.09， 18.09 g·kg-1·d-1， respectively） AMD groups， with 10 rats in each group. The rats in other groups except the blank group were modeled by intraperitoneal injection of p-chlorophenylalanine （PCPA） and administrated with corresponding drugs by gavage， and the blank group received an equal volume of normal saline. The general condition， body mass， and 24 h autonomic activity of each group were observed. The pathological changes of the liver tissue were observed by hematoxylin-eosin（HE）staining and Masson staining. The expression of gamma-aminobutyric acid （GABA）， 5-hydroxytryptamine （5-HT）， epinephrine （EPI）， norepinephrine （NE）， and acetylcholine （ACh） in the liver tissue was detected by enzyme-linked immunosorbent assay. The glutamate （Glu） expression in the liver tissue was detected by the biochemical method. The mRNA levels of biological clock genes Per1， Per2， Cry1， Cry2， Bmal1， and Bmal2 in the liver were determined by Real-time fluorescence quantitative polymerase chain reaction（Real-time PCR）. The protein and mRNA levels of factors in the OX1R/PLCβ-1/PKCα/ERK1/2 signaling pathway in the liver were determined by Western blot and Real-time PCR， respectively. ResultCompared with the blank group， the modeling decreased the body mass （P<0.05， P<0.01） and caused mania and disturbed resting rhythms （P<0.01）， hepatic muscle fiber fracture， and edema with inflammatory cell infiltration. In addition， the modeling decreased the GABA， 5-HT， EPI， NE， and ACh content， increased Glu content （P<0.01）， down-regulated the mRNA levels of Per1， Per2， Cry1， and Cry2 （P<0.01）， up-regulated the mRNA levels of Bmal1 and Bmal2 （P<0.01）， and promoted the expression of OX1R， PLCβ-1， PKCα， and ERK1/2 at both protein and mRNA levels （P<0.01）. Compared with the model group， suvorexant and AMD increased the body mass （P<0.05， P<0.01）， alleviated the mania， and increased the resting time and frequency （P<0.05， P<0.01）. Moreover， the medications elevated the levels of GABA， 5-HT， EPI， NE， and ACh， lowered the Glu level， up-regulated the mRNA levels of Per1， Per2， Cry1， and Cry2 （P<0.05， P<0.01）， down-regulated the mRNA levels of Bmal1 and Bmal2， and inhibited the expression of OX1R， PLCβ-1， PKCα， and ERK1/2 at both mRNA and protein levels （P<0.05， P<0.01）. ConclusionAMD can regulate hepatic neurotransmitters and improve circadian rhythm in insomniac rats by inhibiting the OX1R/PLCβ-1/PKCα/ERK1/2 signaling pathway， and high-dose AMD demonstrated the strongest effect.
ABSTRACT
Objective To evaluate the role of protein kinase Cα( PKCα)?nuclear factor E2?related factor 2 ( Nrf2)?heme oxygenase?1 ( HO?1) signaling pathway on endotoxic shock?induced acute lung injury ( ALI) in rabbits. Methods Thirty healthy male New Zealand white rabbits, aged 2 months, weighing 2?0-2?5 kg, were randomly divided into 3 groups ( n=10 each) using a random number table: normal control group ( group C);ALI group ( group ALI);PKCα inhibitor chelerythrine group ( group CHE) . In group CHE, chelerythrine 8 mg∕kg ( in 0?5 ml of DMSO) was injected intraperitoneally, and 30 min later, LPS 5 mg∕kg ( in 2 ml of normal saline) was injected via the auricular vein to induce ALI in ALI and CHE groups. The rabbits were then sacrificed at 6 h after injection of LPS or normal saline, and the lungs were removed for examination of the pathological changes which were scored and for determination of wet∕dry lung weight ratio ( W∕D ratio) , and the expression of Nrf2 and HO?1 protein and mRNA. Results Compared with group C, the pathological score and W∕D ratio were significantly increased, and the expression of Nrf2 and HO?1 protein and mRNA was up?regulated in ALI and CHE groups. The pathological score and W∕D ratio were significantly higher, and the expression of Nrf2 and HO?1 protein and mRNA was lower in group CHE than in group ALI. Conclusion The PKCα?Nrf2?HO?1 signaling pathway is one of the endogenous protective mechanisms underlying endotoxic shock?induced ALI in rabbits.
ABSTRACT
ObjectiveTo investigate the expression and relevant function of protein kinase C (PKC)-α in kidney of rat with chronic arsenic poisoning.MethodsTotally 60 healthy SD rats of clean grade were randomly divided by body weight into 3 groups:high-dose arsenic exposure group (10.0 mg/kg),low-dose arsenic exposure group (0.4 mg/kg),and control group.The rats were exposed by drinking arsenic solution which was mixed with distilled water.Rats were weighed every 10 days and dose volume of arsenic solution was adjusted.After continuous exposure for 4 months,blood and urinary arsenic were determined.Rat kidneys were taken and stained by Immunohistochemistry SABC.PKC-o positive cells in the kidney were observed and counted,and its average gray value was analyzed with image analysis software (Biomias).ResultsProximal tubules PKC-α-positive cell count [(3.62 ± 1.90),(10.07 ± 3.22)/field],glomerular PKC-α-positive cell count [(3.62 ± 1.90),(10.07 ± 3.22)/field]in high and low arsenic group of SD rat kidney were lower than those of the control group [(60.00 ± 9.63),(18.57 ± 2.71/field,all P ＜ 0.05]; both urinary arsenic level[(7366.62 ± 1086.50),(1744.31 ± 300.12)μg,/L]and blood arsenic level [(31.59 ± 9.24),(16.58 ± 2.08)μg/L] in high-dose and low-dose groups were higher than those of the control group [(18.97 ± 3.58),(18.97 ± 3.58)μg/L,all P ＜ 0.05] ; the average gray values of SD rat kidney proximal tubule,glomerular PKC-o positive cells in high-dose and low-dose groups( 142.79 ± 11.16,122.15 ±5.91 ) were higher than that of the control group (114.33 ± 6.70,all P ＜ 0.05).ConclusionsArsenic can decrease SD rat kidney PKC-α -positive cells.The regulatory function of PKC-o in inhibiting cell apoptosis of kidney of rats with arsenic poisoning is weakened.