|
Objective: liver fibrosis (hepatic fibrosis) liver cell injury caused by a variety of pathogenic factors, a complex pathological process secondary to inflammation of the liver and liver tissue to repair itself. This process led to the synthesis and degradation of extracellular matrix (extracellular matrix, ECM) imbalance, causing ECM excessive deposition of the formation of liver fibrosis in the liver tissue, and even then cirrhosis. In the process of liver fibrosis, cytokine plays a key role. Hepatic stellate cells (hepatic stellate cell, HSC) is a major cell secretion of various cytokines and produce ECM HSC activation, proliferation is a key link of liver fibrosis. HSC proliferation, activation and synthesis of ECM liver fibrosis, a variety of cytokines such as transforming growth factor-β1 (transforming growth factor-β1, TGF-β1), insulin-like growth factor -1 (insulin-like growth factor-1 , IGF-1) plays an important role. The physiological state of the ECM synthesis and degradation in a dynamic balance, in order to maintain normal liver tissue structure and function. However, damage the liver of a variety of pathogenic factors, to produce the inflammatory response, inflammatory cytokines HSC changed from a stationary state to the activated state, the secretion of various cytokines to promote HSC activation and proliferation, while generating the ECM, to make it in the liver the deposition of the formation of fibrosis and even then hardened. IGF-1 is a regulator of cell growth, differentiation, proliferation and metabolism of a polypeptide growth factors. In vitro studies have found that IGF-1 can promote HSC activation, proliferation and collagen synthesis, and promote the formation of liver fibrosis. However, IGF-1 in the process of liver fibrosis is still not very clear. Liver fibrosis in chronic liver disease progression to cirrhosis only way. Cirrhosis of the liver is the end stage of chronic liver disease, there may be a variety of complications, causing great pain to the patient. The treatment of liver fibrosis control the cirrhosis key. It has now been confirmed interferon and nucleos (t) ide analogs, by inhibition of viral replication, reduce inflammation, inhibit fibrosis. However, antifibrotic clinical types of western medicine and the limited efficacy. Chinese medicine with its multi-channel, multi-target therapy regulatory role in the antifibrotic a unique advantage. The Fuzhenghuayu party fermentation cordyceps Salvia, pine pollen, peach, Gynostemma Chinese medicine believes that its stasis, the Yijing Yanggan role of modern research that effective reverse liver fibrosis. Therefore, we established a rat model of hepatic fibrosis induced by carbon tetrachloride (CCL4), to investigate the IGF-1 dynamic changes in experimental liver fibrosis, while intervention Fuzhenghuayu party to observe the righting of stasis square hepatic fibrosis and its impact on the role of IGF-1, to provide a theoretical basis for seeking effective antifibrotic drugs. Method: The first part of the experiment is divided into two parts: IGF-1 in rat liver fibrosis spontaneous dynamic changes in the reversal process selected 66 male wistar rats were randomly divided into control group, model group. Control group were injected with saline, the model group were given 30? L4 olive oil intraperitoneal injection, are 2 times per week for 8 weeks. Started in modeling the first 4 weeks, every two weeks, randomly taken from the model rats were sacrificed 6, the control group 3 were observed to 14 weekend. The second part of experimental liver fibrosis and liver tissue IGF-1: Fuzhenghuayu side selected 57 male wistar rats were randomly divided into a control group, model group and Fuzhenghuayu of party intervention group. Control group, intraperitoneal injection of saline, 2 times per week for 8 weeks; model group were given 30? L4 olive oil intraperitoneal injection twice a week for 8 weeks; intervention group modeling approach with model group, while giving Fuzhenghuayu side liquid medicine orally, once daily for 8 weeks. Randomly taken from the model group and intervention on the weekend of the first 4,6,8 rats were sacrificed 6, the control group were sacrificed 3. The rats were sacrificed, specimens of blood and liver tissue samples of serum alanine aminotransferase enzyme (ALT), aspartate aminotransferase enzyme (AST); hematoxylin - eosin (HE) and Masson staining observed liver histopathological changes; alkali The immunohistochemistry staining liver tissue α-smooth muscle actin (α-smooth muscle actin, α-SMA), the expression of the IGF-1 protein; RT-PCR was used to detect the expression of IGF-1 mRNA; hydrolysis assay liver hydroxyproline content (hydroxyproline, Hyp). Results: The first part: of IGF-1 in rat liver fibrosis spontaneous reversal process of dynamic change a serological indicator model of serum ALT, AST increased gradually at 4, 6 and 8 weeks, 10 weeks began to decrease, still higher than in the first 14 weeks. The model rats each time period, ALT and AST was higher than that in the control group. Liver histopathological changes in HE and Masson staining: control group normal lobular architecture rat liver tissue, liver cell cords running neat, no inflammatory cell infiltration in the vessel wall, the portal area visible to the presence of a small amount of fibrous connective tissue; model group in the first four weeks of the liver tissue, hepatic lobule structure is damaged, the visible liver cell degeneration, necrosis, inflammatory cells infiltration; six weeks in the fourth week on the basis of visible liver tissue of hepatic steatosis, portal area gradually visible a lot of fibrous tissue hyperplasia; 8 weeks liver steatosis disappeared, showing that a large number of fiber cord stark pseudolobules. The late inflammatory cells gradually reduce, reduce fibrous connective tissue. 3 liver tissue α-SMA and IGF-1 protein expression to 3.1 liver tissue protein expression of α-SMA: immunohistochemical staining α-SMA in each period of the model group compared with the control group were significantly increased. The control group only α-SMA expression in the vascular wall; With the modeling time extension, model group, α-SMA expression increased interval were positive expression in the vascular wall, portal area and fiber gradually reduced after 10 weeks. 3.2 liver tissue IGF-1 protein expression: Immunohistochemical staining showed that the control group to express a small amount of IGF-1 protein in rat liver cytoplasm; model group 4,6,8 weeks IGF-1 expression gradually increased, IGF-1 in the liver cytoplasm and cell membrane expression in late relieve inflammation and fibrosis of the liver tissue, decreased gradually. 4 liver tissue IGF-1 mRNA expression by RT-PCR display model group each time period liver tissue IGF-1 mRNA expression than in the control group was significantly increased, the first eight weeks of IGF-1 mRNA expression to the highest, and later as CCL4 applications Stop and improved liver fibrosis, IGF-1 mRNA expression was gradually reduced. 5 liver tissue Hyp detection model group as CCL4 the application, hepatic Hyp content gradually increased, peaked to eight weeks after in CCL4 the disable and liver fibrosis reversal, Hyp decreased gradually. Hyp content of the liver tissue of the model group at each time point were higher. 6 liver of Hyp content in liver tissue α-SMA and IGF-1 protein expression in the relationship between IGF-1 and α-SMA protein expression in liver tissue Hyp content between straight-line relationship, the correlation coefficients were 0.953 and 0.944 (P < 0.01). The second part: Fuzhenghuayu square experimental liver fibrosis and liver tissue IGF-1 serum markers the model group Fuzhenghuayu intervention serum ALT, AST 4, 6 and 8 weeks compared The control group was significantly higher in the intervention group rat serum ALT, AST model group was significantly lower than in the corresponding period. 2 liver histopathological changes in HE and Masson staining rat liver tissue: the control group lobular architecture normal, liver cell cords neatly, a small amount of fibrous connective tissue in the vessel wall and the portal area visible; lobular architecture of the model rats be destroyed, the visible liver cell degeneration, necrosis, a large number of inflammatory cell infiltration, gradually increased with the progression of fibrosis, a large number of fibrous tissue hyperplasia, formation pseudolobules; inflammation and fibrosis in rat liver tissue intervention group compared with the model group, the same time period The extent of significantly reduced. Liver tissue α-SMA and IGF-1 protein expression 3.1 liver tissue protein expression of α-SMA: Immunohistochemical staining showed that the control group only α-SMA expression in the vascular wall; extension of time with the modeling, model group, α-SMA expression gradually increased, and have a positive expression in the vascular wall, portal area and fibrous septa. Intervention group, α-SMA expression of the same period compared with the model group were significantly lower. 3.2 liver tissue IGF-1 protein expression: Immunohistochemical staining showed that the control group to express a small amount of IGF-1 protein in rat liver cytoplasm; model group, the expression of IGF-1 compared with the control group increased; intervention group with the same period model group, the expression of IGF-1 Compared were significantly reduced. The 4 liver tissue IGF-1 mRNA expression by RT-PCR liver tissue of the model group and the intervention group IGF-1 mRNA expression compared with the control group significantly increased, consistent trend with IGF-1 protein expression; intervention group IGF-1 mRNA in various the time periods expression were significantly lower than the model group. Hyp content of the 5 liver tissue Hyp detection model group and the intervention group and the control group were significantly increased, Hyp content of the intervention group were significantly lower than in the expression of each period the model group. Conclusion: rat CCL4 intraperitoneal injection of 8 weeks, can induce the typical liver fibrosis spontaneous reversal model, the pathological features consistent with the characteristics of human liver fibrosis. 2 IGF-1 is closely related to the degree of liver fibrosis and inflammation necrosis, may have a role to promote liver fibrosis. 3 IGF-1 protein expression of α-SMA in liver tissues trends, and IGF-1 protein expression of α-SMA and Hyp content linear relationship, IGF-1 may promote liver by promoting HSC activation and proliferation, fibrosis. 4 Fuzhenghuayu able to significantly inhibit liver inflammation and fibrosis and improve liver function. The 5 Fuzhenghuayu party may through inhibition of IGF-1, α-SMA protein, to reduce liver tissue Hyp content, inhibition of HSC activation, proliferation, and thus play the role of anti-liver fibrosis.
|