Farnesoid X receptor up-regulates thyrotropin embryonic factor and attenuates pathological injury of Con A-induced hepatitis

AIM：To observe how farnesoid X receptor (FXR) functioned in concanavalin A (Con A) -induced hepatitis (CIH) and the regulation of FXR-thyrotropin embryonic factor (TEF) pathway. METHODS：C57BL/6 mice were injected with Con A to induce hepatitis. The expression of FXR and TEF in the liver specimens was determined by qRT-PCR and Western blotting. The concentrations of serum ALT/AST and inflammatory cytokines IFN-γ, TNF-α, IL-4 and IL-2 in the blood samples were tested after Con A injection. RESULTS：FXR was down-regulated in CIH mice. TEF was up-regulated when FXR was activated by chenodeoxycholic acid (CDCA). Activation of FXR reduced the levels of aminotransferases and inflammatory cytokines IFN-γ, TNF-α, IL-4 and IL-2 in the CIH mice induced by Con A injection. CONCLUSION：FXR activation attenuates CIH mouse liver injury and reduces inflammatory cytokines. FXR activation results in TEF up-regulation. The FXR-TEF pathway may play a protective role in autoimmune hepatitis.     

Effect of portal vein transplantation with allogeneic adipose tissue-derived mesenchymal stem cells on liver fibrosis in SD rats

AIM: To explore the influence of adipose tissue-derived mesenchymal stem cells (ADMSCs) transplantation on hepatic fibrosis in rats. METHODS: ADMSCs from abdominal lipid tissues were extracted, cultured and passaged. The hepatic fibrosis rat model was built up and randomly divided into 3 groups: hepatic cirrhosis group (n=14); portal vein transplantation group (n=11) and caudal vein transplantation group (n=14). Computer tomography(CT) perfusion index, histological scores and microvessel density were detected and compared after transplantation of ADMSCs among the 3 groups. RESULTS: After transplantation of ADMSCs, the total hepatic blood perfusion, especially portal vein perfusion, significantly increased in portal vein transplantation group determined by CT perfusion scan (P<0.05), but slightly increased in caudal vein transplantation group. The histological scores showed significant alleviation of fibrosis evidence in portal vein transplantation group, and slightly change of adipose degeneration in caudal vein transplantation group. Microvessel density decreased significantly in portal vein transplantation group as compared to the other 2 groups (P<0.05). CONCLUSION: Transplantation of ADMSCs greatly helps the alleviation of hepatic fibrosis. Portal vein transplantation benefits more than caudal vein transplantation.     

Minimally modified low-density lipoprotein up-regulates endothelin receptors in mouse mesenteric artery through p38 MAPK pathway

AIMTo investigate whether minimally modified low-density lipoprotein (mmLDL) affects the quantity and activity of endothelin （ET） type A (ET_A) and type B (ET_B) receptors in mouse mesenteric artery by activating p38 mitogen-activated protein kinase (MAPK) inflammatory pathway. METHODSThe KM mice were divided into normal saline (NS) group (injection of NS via caudal vein), mmLDL group (injection of mmLDL via caudal vein), LDL group (injection of LDL via caudal vein), mmLDL+SB 203580 group (injection of mmLDL via caudal vein and intraperitoneal injection of p38 MAPK pathway specific inhibitor SB 203580) and mmLDL+DMSO group (injection of mmLDL via caudal vein and intraperitoneal injection of DMSO). Mesenteric artery ring segment vasoconstriction dose-response curves affected by sarafotoxin 6c (S6c) and ET-1 were recorded by the myography system. The mRNA levels of ET_B receptor, ET_A receptor and interleukin-6 (IL-6) were detected by RT-qPCR. The protein levels of ET_B receptor, ET_A receptor, IL-6, p38 MAPK, p-p38 MAPK, NF-κB and p-NF-κB were determined by Western blot. The serum concentration of IL-6 was measured by ELISA. RESULTSThe contractile responses of the blood vessel segments to S6c and ET-1 were significantly increased by mmLDL (P<0.01). The mRNA and protein expression levels of ET_A receptor, ET_B receptor, and IL-6 significantly increased (P<0.01). The protein levels of p-p38 MAPK and p-NF-κB were significantly increased (P<0.01). The serum level of IL-6 was significantly increased (P<0.01). These effects of mmLDL were inhibited by p38 MAPK inhibitor SB 203580. CONCLUSION mmLDL increses the serum concentration of IL-6, up-regulates the expression of IL-6, ET_A receptor and ET_B receptor in mouse mesenteric artery, and enhances the vasoconstriction function medi?ated by ET_A and ET_B receptors, which is related to the activation of p38 MAPK inflammatory pathway and downstream NF-κB pathway.     

Effect of TLR2/4 ligand-enhanced non-specific liver inflammation on autoimmune hepatitis in BALB/c mice

AIM: To explore the role of non-specific liver inflammation in inducing autoimmune hepatitis (AIH) in BALB/c mice.METHODS: The plasmids pCYP2D6, pcDNA3.1 and psTLR2/4 were administered by tail vein injection. The carbon tetrachloride (CCl₄) was injected in the abdominal cavity. The autoimmune response was measured by ELISA. The liver inflammation was observed by HE staining. The liver fibrosis was evaluated by Sirius red staining. RESULTS: CCl₄ induced non-specific liver inflammation in the BALB/c mice, and TLR2/4 ligand enhanced the inflammatory responses. After the repeated injection of CCl₄ stopped, the non-specific liver inflammation disappeared, but CCl₄ promoted autoimmune response, autoimmune hepatitis and liver fibrosis induced by mimetic antigen human CYP2D6, and TLR2/4 ligand enhanced these changes. CONCLUSION: TLR2/4-amplified liver non-specific inflammation may play an important role in the initiation and progression of autoimmune hepatitis in BALB/c mice.     

Effect of HOCs implantation on protein expression of TGF-β/Smad signaling pathway in liver tissue of hepatic fibrosis rats

AIM: To investigate the effect of hepatic oval cells (HOCs) on the protein expression of TGF-β/Smad signaling pathway in the liver tissue of hepatic fibrosis rats.METHODS: The SD rat models of liver fibrosis were made by treating with carbon tetrachloride and combined factors. The HOCs was isolated from the model rats. HOCs suspension (0.5 mL at a density of 1×1012cells/L) were transplanted via portal vein into the hepatic fibrosis rats at 8th week and observed continuously for 30 days. Meanwhile, WuLing capsules were used for positive control. The blood samples were collected through trail vein at 8th day, 15th day, 23th day and 30th day after transplantation of HOCs. The levels of aspartate aminotransferase (AST) and alamine aminotransferase (ALT) in serum were determined by enzyme method. The morphological changes of hepatic tissues were observed under microscope with HE and Musson staining. The protein levels of collagen type I (Col-Ⅰ), extracellular-signal regulated protein kinase (ERK), phosphory-lation extracellular regulatedprotein kinases (p-ERK), TGF-β receptor type Ⅰ (TβRⅠ), TGF-β receptor type Ⅱ (TβRⅡ), mothers against decapentaplegic homolog 2/3 (Smad 2/3) and mothers against decapentaplegic homolog (Smad 7) were assessed in liver tissues by Western blotting. RESULTS: In HOCs and WuLing capsules treated groups, the levels of ALT and AST decreased significantly at 15th day, 23th day and 30th day after the transplantation of HOC. The damage degree of hepatic fiber hyperplasia of the liver histological structure reduced notably. The expression levels of Col-Ⅰ, ERK, p-ERK, TβRⅠ and TβRⅡ in liver tissues of hepatic fibrosis rats were down-regulated obviously while the expression of Smad 7 increased significantly.CONCLUSION: The implantation of HOCs prevents the progress of liver fibrosis in rats. The mechanism of action is to inhibit the protein expression of p-ERK, TβRⅠ, TβR Ⅱ for TGF-β/Smad signaling pathway of liver tissue.     

Responsiveness of the T-lymphocyte in peripheral blood from asthmatic patients to VIP

AIM: To investigate the functional state of vasoactive intestinal peptide (VIP) receptor on T-lymphocyte (T-cell)from asthmatics. METHODS: T-cells were purified from peripheral blood of asthmatics in remission and control subjects. Proliferation of T-cells was measured by [³H][³H]-TdR incorporation rate. The cAMP level in T-cells was assayed with radioimmunoassay method. We observed the influence of VIP on Con A-induced proliferation of T-cells, and cAMP level in T-cells in asthmatic and control groups. RESULTS: There was no difference in Con A-induced proliferation of T-cells between the two groups (P＞0.05). VIP, however, could inhibit the Con A-induced proliferation of T-cells from control subjects more significantly than that from asthmatics (P＜0.01). There was no difference in cAMP level in the unstimulated T-cells between the two groups (P＞0.05). The cAMP level in T-cells, however, increased more significantly in the control group than that in the asthmatic group after the treatment of VIP or NaF (P＜0.01,respectively). Adenylcyclase stimulator (forskolin)-treatment had no effect on the cAMP level in T-cells from the two groups (P＞0.05). CONCLUSION: Inhibition effect of VIP on Con A-induced proliferation of T-cells was less in asthmatics than in control subjects, which may be related to insufficiency of Gs α coupled VIP receptor on T-lymphocytes in asthmatics.     

Transplantation of bone marrow mesenchymal stem cells improves motor function in a DMD mouse model

AIM: To observe amelioration of motor function in a Duchenne muscular dystrophy (DMD) mouse model (dko mice) after transplantation of bone marrow mesenchymal stem cells (MSCs). METHODS: Passage fifth MSCs cultured in vitro were transplanted into dko mice by tail vein, motor functions of experimental mice and matched control mice, including traction, rotating rods, rotated wheel, upside down, turning over and walking (all were recorded by Sony digital camera) were tested 15 weeks after transplantation. The fluorescent expression of dystrophin and utrophin in gastrocnemius muscle tissue of dko mice was detected by SABC-Cy3, and average optical density of positive fibers was calculated. RESULTS: MSCs grew in colony over passage third, and there was low immunologic reaction by vein transplantation. There was dystrophin and utrophin fluorescent expression in sarcolemma of dko mice 15 weeks after transplantation, but no any fluorescent expression in controls. There was significant difference in fluorescent average optical density of positive fibers between two groups (P＜0.05). Amelioration of motor functions in dko mice was found 15 weeks after MSCs transplantation compared with the control mice (P＜0.05). CONCLUSION: Transplantation of MSCs ameliorates the positive and passive motor functions of dko mice.     

Transplantation of mouse fetus-derived MSCs alleviates physical fatigue in mouse

AIM:To observe the effect of mouse fetus-derived mesenchymal stem cell(MSC) transplantation on alleviating fatigue in mice. METHODS:MSCs were derived from the mouse fetus at 13.5 d. The 6-month-old BALB/c mice were randomly divided into non-swimming group, swimming control group and swimming+MSCs group. The mice in swimming+MSCs group were injected with fetus-derived MSCs through the tail vein, while the mice in non-swimming group and swimming control group were injected with equal volume of normal saline. The anti-fatigue activity was evaluated using a weight-loaded swimming test, along with the determination of blood urea nitrogen(BUN), blood lactic acid(BLA),and glycogen in the hepatic and muscular tissues 24 h later. The antioxidant activity was evaluated by determining the activity of superoxide dismutase(SOD) and the content of malondialdehyde(MDA). The cardiac functions were measured by echocardiography. RESULTS:Primary cultured cells were spindle-shaped in scattered colony. The cells in fifth passage grew parallelly or swirlingly. The cells expressed CD44 and CD105, but not CD34 or CD45. Inducing experiments showed that the cells differentiated into osteoblasts or adipocytes. Transplantation of MSCs alleviated fatigue in mice. In swimming+MSCs group, weight-loaded swimming time was longer, the levels of BUN, BLA, hepatic and muscular MDA were lower, and the levels of glycogen and SOD in the hepatic and muscular tissues were higher than those in control group. Stroke volume, cardiac output and left ventricular diastolic volume in swimming+MSCs group increased, indicating that the cardiac functions were enhanced. CONCLUSION: Transplantation of mouse fetus-derived MSCs alleviates physical fatigue.     

Chronic alcohol intake-induced epithelial-mesenchymal transition contri-butes to hepatic fibrogenesis in mice

AIM：To evaluate the effect of chronic alcohol intake on the histopathological changes of the liver and to determine the contribution of epithelial-mesenchymal transition (EMT) to hepatic fibrogenesis. METHODS：Thirty male C57BL/6 mice were randomly divided into 3 groups as following: the mice in control group was given (ig) water; the mice in low-dose alcohol group (2.0 g·kg -1·d -1) and high-dose alcohol group (4.0 g·kg -1·d -1) were given (ig) alcohol for 5 months. Alcohol-induced histopathological changes of the liver or development of hepatic fibrosis were evaluated using the histological methods with HE and Masson trichrome staining. The apoptosis of the liver was detected by TUNEL fluorometric staining (counterstained with DAPI). The activity of serum alanine aminotransferase (ALT) and aspartate aminotransferase (AST) was measured by an automated biochemical analyzer. The expression of fibroblast-specific protein 1 (FSP-1), α-smooth muscle actin (α-SMA) and E-cadherin in the hepatic tissues was detected by immunofluorescence examination. The protein levels of E-cadherin, α-SMA, FSP-1, transforming growth factor β 1 (TGF-β 1) and hypoxia-inducible factor 1α (HIF-1α) were analyzed by Western blotting. RESULTS：Compared with control, the activity of serum ALT and AST, and apoptotic index of liver tissues were increased in the mice treated with alcohol for 5 months. The histopathological changes of the livers in the mice of low-dose alcohol group included steatosis and mild liver fibrosis, while severe liver fibrosis was observed in the high-dose alcohol-treated mice. Chronic alcohol consumption induced the increase in malondialdehyde (MDA) level, and the decreases in the activity of superoxide dismutase (SOD) and catalase (CAT) in the livers. It also reduced E-cadherin expression and increased α-SMA expression. FSP-1 immunostaining and albumn immunostaining positive cells were co-localized in the hepatocytes of low-dose alcohol group, but only FSP-1 positive hepatocytes were observed in high-dose alcohol group. Chronic alcohol consumption decreased E-cadherin expression and increased α-SMA, FSP-1, TGF-β 1 and HIF-1α expression in a dose-dependent manner, but the HIF-1α expression was not altered between the 2 alcohol-treated groups. CONCLUSION：Chronic alcohol intake induces the progression of hepatic fibrosis. Some fibroblasts derive from hepatocytes in liver fibrosis via EMT. The underlying mechanism is associated with the changes of the redox state, and increased TGF-β 1 generation and HIF-1α expression.     

Riboflavin preconditioning alleviates hepatic ischemia/reperfusion injury in rats

AIM: To explore the protective effect of riboflavin preconditioning on hepatic ischemia/reperfusion injury in rats. METHODS: Twenty-four Sprague-Dawley rats wererandomly divided into 3 groups (n=8): sham group, ischemia/reperfusion (I/R) group and riboflavin preconditioning (R+I/R) group. The rats in sham group and I/R group received a standard chow,while the rats in R+I/R group received a chow supplemented with riboflavin. After 4 weeks, portal vein and hepatic artery supplying the middle and left hepatic lobes were clamped with a traumatic vascular clip for induction of partial hepatic ischemia in the rats in I/R group and R+I/R group. After 1 h of ischemia, 1 h of reperfusion was conducted by removal of the clip. The activity of alanine aminotransferase (ALT) and aspartate aminotransferase (AST) in serum,the activity of superoxide dismutase (SOD) and the level of malondialdehyde (MDA) in serum and liver were measured. Western blotting was employed to examine the protein expression of heme oxygenase-1(HO-1) in the liver. RESULTS: The results showed that ischemia/reperfusion injury markedly increased the activity of AST and ALT in serum, decreased the activity of SOD, and elevated the level of MDA and the activity of HO-1 in the liver as compared with sham group (P<0.01). The riboflavin pretreatment significantly decreased the activity of AST and ALT in serum, increased the activity of SOD and decreased the levels of MDA in serum and liver as compared with I/R group (P<0.01). In addition, the protein expression of HO-1 and the activity of HO-1 were elevated in R+I/R group (P<0.01). Cytoplasmic vacuolation and swelling of the hepatocytes were observed in I/R group. Treatment with riboflavin markedly alleviated the changes of liver structure. CONCLUSION: Riboflavin preconditioning has protective effect on hepatic ischemia/reperfusion injury. The mechanism may be correlated with enhancing the anti-oxidation and alleviating the reaction of lipid peroxidation.     

Intervention of Lipo-PGE1 on liver blood perfusion by different time and medication: a empirical study

AIM: To explore the effect and mechanism of liposome prostaglandin E₁(Lipo-PGE₁) on liver blood perfusion by different time and medication.METHODS: Twelve healthy adult dogs were injected with Lipo-PGE₁₁ μg/kg via left small saphenous vein at speed of 0.05 μg·kg^-1·min^-1.Liver computed tomography perfusion imaging (CTPI) was performed on 0,5,15 and 30 min,and the value of hepatic arterial perfusion (HAP),portal vein perfusion (PVP) and total liver perfusion (TLP) among groups were compared.The impacts of Lipo-PGE₁ on liver haemodynamics at different time were investigated.Twenty-four health dogs were randomly divided into four groups: control group,peripheral vein group,hepatic artery group and superior mesenteric artery group.Liver CTPI was performed at 5 min after 1 μg/kg Lipo-PGE₁ administration in those groups.The values of HAP,PVP and TLP were compared and effects of Lipo-PGE₁ on liver blood flow by different medication were observed.RESULTS: The values of liver perfusion (mL·min^-1·mL^-1) at 0,5,15 and 30 min after 1 μg/kg Lipo-PGE₁ administration via vein were as follows: HAP: 0.22 ±0.65,0.24±0.65,0.22±0.69,0.22±0.06;PVP: 1.22±0.40,1.88±0.59,1.55±0.55,1.29 ±0.57;TLP: 1.44±0.42,2.12±0.61,1.77±0.56,1.51±0.58,respectively.No significant difference in HAP among groups was observed,but in PVP and TLP,significant differences (F=3.812,P<0.05;F=3.805,P<0.05) among groups were found.The values of PVP and TLP were most obviously increased at 5 min,and the values of PVP and TLP were still on the high level at 15 min and 30 min.The values of liver perfusion (mL·min^-1·mL^-1) by different medication were as fellows: HAP: 0.22±0.06,0.24±0.06,0.31±0.07,0.26±0.05;PVP: 1.28±0.38,2.33±0.41,2.37±0.55,2.83±0.94;TLP: 1.50±0.40,2.57±0.42,2.67± 0.58,3.09±0.94,respectively.No significant difference in HAP among groups (F=2.248,P>0.05) was found,but in PVP and TLP group,significant differences (F=6.892,P<0.01;F=7.802,P<0.01) among groups were observed.In addition,superior mesenteric artery group showed higher value of PVP and TLP than other methods.CONCLUSION: Lipo-PGE₁ obviously increases liver blood perfusion,especially for portal vein perfusion.Interventional technology provides an effective pathway to improve hepatic perfusion.     

Effect of camptothecin on the activation,proliferation and cell cycle distribution of the mouse T lymphocytes in vitro

AIM: To study the effects of camptothecin (CPT) on the activation, proliferation and cell-cycle distribution of the mouse T lymphocytes stimulated by concanvalin A (ConA) in vitro. METHODS: A model of T cell activation and proliferation was established by stimulated the cells with Con A. T cells were treated with different concentrations of CPT. The expression of CD69, the early marker of CD3+ T cell activation, was measured by FACS. The proliferation index was determined by carboxyl fluorescin diacetate succinmidyl ester by flow cytometry. The cell-cycle distribution was analyzed by propidium iodide staining. RESULTS: After stimulation with Con A for 6 h, the activation rate of CD69+ T cell in Con A group was (58.88±0.55)%. The percentages of CD69 positive cells were (55.48±0.98)%, (54.67±1.05)%, (50.40±0.82)%, (42.47±1.32)%, correspond to the treatments with different concentrations of CPT (10 nmol/L, 20 nmol/L, 50 nmol/L, 100 nmol/L), respectively. After 48 h treatment with Con A, the proliferation index in different concentrations of CPT treatment (10 nmol/L, 20 nmol/L, 50 nmol/L and 100 nmol/L) exerted a definite inhibitory effect on the proliferation (P<0.01). Moreover, the cell-cycle distribution analysis showed that apoptosis peak was observed in different concentrations of CPT treatment after 48 h cultured with Con A. CONCLUSION: CPT significantly inhibits the early stages of the Con A-induced T cell activation and proliferation, and detents the T lymphocytes in G₀/G₁ phase.     

Effect of hepatitis virus B protein on the functions of PBMCs isolated from patients with chronic HBV infection

AIM: To study the effect of hepatitis virus B proteins on peripheral blood mononuclear cells (PBMCs) from patients among various types of chronic hepatitis B virus (HBV) infection.METHODS: 80 patients of various types of chronic HBV infection were observed, including 40 HBeAg positive with abnormal alanine aminotransferase (ALT) (A group), 20 HBeAg positive with persistent normal ALT(B group), 20 HBeAg and HBV-DNA negative with persistent normal ALT level(C group). IL-10, IFN-γ in CD8+CD28+T cells, after stimulation with PHA, HBeAg and HBcAg for 48 h, were inspected respectively in PBMCs.RESULTS: IFN-γ was significantly lower in HBeAg positive patients. IL-10 was significantly higher in HBeAg positive with normal ALT. CD8+CD28+T were significantly lower than others. CONCLUSION: In HBeAg positive group, secretion of cytotoxic T lymphocyte (CTL) and Th1 type cellular immunologic reaction is decreased, Th2 type cellular immunologic reaction is enhanced.     

Non-viral vector-mediated high efficiency expression of manganese superoxide dismutase gene in mouse liver

AIM: To investigate the transfection efficiency of mouse liver with non-viral vector containing manganese superoxide dismutase (Mn-SOD) gene. METHODS: The eukaryotic expression vector, gWiz/Mn-SOD, encoding human manganese superoxide dismutase was constructed. The plasmids of gWiz/Mn-SOD were mixed with cationic lipids, followed by injection into mice via branch of superior mesenteric vein, to induce Mn-SOD over-expression in murine liver detected by RT-PCR, Western blotting, SOD activity and immunohistochemical staining. RESULTS: gWiz/Mn-SOD transfection resulted in the obvious expression of exogenous Mn-SOD mRNA and protein in hepatic tissues at 8 hours after injection, and elevated mitochondria SOD activity 8.4 times in transfected hepatocytes than that in non-transfected cells at 72 hours after injection. It was showed that nearly 70% of mouse hepatocytes was obviously Mn-SOD positive after transfection. CONCLUSION: High expression efficiency of Mn-SOD gene in mouse liver is achieved safely, by injection of gWiz/Mn-SOD and cationic lipid mixture into branch of superior mesenteric vein.     

Hepatocyte protection of ethyl pyruvate in septic mice

AIM:To study the protective effect of ethyl pyruvate (EP) on hepatocytes in septic mice. METHODS:The cecal ligation-perforation was made in mice as septic model. Ringers ethyl pyruvate solution (REPS) and Ringers lactic solution (RLS) were used to resuscitate septic mice. Anti-oxidative capacity of hepatic tissue and liver function were detected in different groups. RESULTS:Anti-oxidative capacity in septic mice was significantly lower than that in sham group (P<0.01). EP promoted the anti-oxidative capacity of hepatic tissue in septic mice. Malondialdehyde level was lower in REPS group than that in RLS group ［(48.18±5.98) μmol·g-1 protein vs (78.34±11.16) μmol·g-1 protein］, superoxide dismutase ［(5.19±1.41)103 U/g protein vs (3.20±1.08)103 U/g protein］ and total anti-oxidative capacity ［(7.02±1.79)103 U/g protein vs (4.77±1.35)103 U/g protein］ level were higher in REPS group than those in RLS group (P<0.01). Alanine aminotransferase in REPS group were lower than that in RLS group ［(210.06±23.36) U vs (458.86±51.55) U, P<0.01］. CONCLUSION:Ethyl pyruvate is an effective anti-oxidant in septic mice, which significantly increases the anti-oxidative capacity in hepatic tissue and ameliorates liver function.     

Role of IL-23/IL-17 inflammatory axis in the pathogenesis of psoriasis-like lesions induced by imiquimod in mice

AIM: To observe the dynamic changes of IL-23/IL-17 inflammatory axis in psoriasis-like lesions of mice induced by imiquimod (IMQ).METHODS: BALB/c female mice were randomly divided into control group and IMQ group. The morphological changes of lesional skin in mice were evaluated according to the psoriasis area and severity index (PASI) and HE staining. cytokine antibody chips were used to determine the cytokine changes in serum and lesions. The mRNA and protein expression of cytokines were analyzed by cytometric bead array, real-time PCR and Western blotting. Moreover, the changes of cellular constituents in the peripheral blood and splenic cells of mice were detected by flow cytometry.RESULTS: Typical psoriasis-like skin lesions, such as red scaly skin plaques, caused by topical IMQ showed a parabolic dynamic change. There was a dynamic increase in proinflammatory cytokines of the IL-23/IL-17 axis in IMQ-treated skin. IMQ application resulted in elevated expression of cytokines related with IL-23/IL-17 inflammatory axis,Th1-type cytokines,Th2-type cytokines and Treg-type cytokines at day 4. IMQ-treated BALB/c mice showed an increased pericentage of dentric cells in peripheral blood and spleen compared with control animals. Percentages of Th17 and Treg in IMQ-treated mice were increased by 3~4 times and twice as compared with control mice, respectively.CONCLUSION: The skin lesions, histopathological features and cytokine changes in mice induced by IMQ are similar to human psoriasis, which are suitable for investigating the pathogenesis of psoriasis as a psoriasis-like model. IL-23/IL-17 axis is involved in the formation of psoriasis-like skin lesions in mice induced by IMQ and presents a dynamic change. Besides, Th1 cell-mediated inflammatory response is also activated in the formation of lesional skin, accompanied by the increase expression of Th2 and Treg cytokines in a feedback mechanism.     

Inhibitory effects of Angelicae sinensis and sodium ferulate on acute inflammatory liver injury and expression of ICAM-1 and E-selectin in mice

AIM: To explore the effects of Angelicae sinensis preparation and sodium ferulate on inflammatory liver injury induced by lipopolysaccharide (LPS) and their molecule mechanism. METHODS: ICR mice were divided into five groups: Angelicae sinensis group, sodium ferulate group, dexamethasone group, inflammation control group and normal group. The model of inflammatory injury in mice was set up by tail vein injection with the bacillus calmette-guerin (BCG) and LPS respectively prior and posterior to administration of those tested drugs. The tested drugs (the preparations of Angelica sinensis, sodium ferulate and dexamethasone) and normal saline were given respectively to the corresponding group. The pathological observation of the liver tissues in mice was made for the intensity of inflammatory liver injuries. The immunohistochemical detection and comparison were performed for the expression of ICAM-1 and E-selectin proteins in the liver tissues in those mice. RESULTS: The intensities of liver inflammatory injuries in mice from drug-treated groups were obviously lighter than that from the inflammation control group (P<0.01). The expressions of ICAM-1 and E-selectin proteins in the liver tissues of mice from the inflammation control group were not only significantly higher than that in normal control, but also obviously higher than that from drug-treated groups (P<0.01). CONCLUSIONS: Angelica sinensis and sodium ferulate alleviate inflammatory liver injury induced by injection of LPS. Their suppressive effects on inflammatory liver injury may be associated with the decrease in the expression of ICAM-1 and E-selectin proteins.     

Activation of PPARs is involved in effect of bezafibrate on diabetic hepatopathy in mice

AIM: To investigate the effects of bezafibrate (BEZ) on diabetic hepatopathy in mice. METHODS: Diabetic hepatopathy model was established by a long-high-energy diet combined with streptozotocin (40 mg·kg^-1·d^-1× 5 d, ip), and then bezafibrate (75 mg·kg^-1·d^-1, ig) was supplemented for 4 weeks. Fasting blood glucose (FBG) was detected every week. The structure of liver was observed by HE staining, and the liver function was measured by observing the levels of alanine aminotransferase (ALT) and aspartate aminotransferase (AST). Total cholesterol (TC), triglyceride (TG), insulin and HbA1c were determined by commercial kits, and then HOMA insulin resistance index (HOMA-IR) was calculated. The expression of peroxisome proliferator-activated receports (PPARs) at mRNA and protein levels was determined by RT-qPCR and Western blot, respectively. RESULTS: After 7 days of treatment with streptozotocin, the FBG level of the mice exceeded 11.1 mmol/L. At the end of the experiment, the structural deformation of the hepatocytes (containing abundant fat vacuoles, infiltrated by inflammatory cells, etc.) was observed, with the increases in insulin, HbA1c, HOMA-IR, TC, TG, ALT and AST in diabetic mice (P<0.01). Meanwhile, the expression of PPARα, PPARβ and PPARγ at mRNA and protein levels was decreased (P<0.01). Bezafibrate treatment markedly attenuated the structural and functional damages of the liver in the diabetic mice (P<0.01), and reduced the levels of FBG, insulin, HbA1c, HOMA-IR, TC and TG (P<0.05). Bezafibrate also up-regulated the expression of PPARα, PPARβ and PPARγ (P<0.01). CONCLUSION: Bezafibrate attenuates hepatic injury in diabetic mice via the activation of PPARs-related signaling pathway.     

Ultrastructural characteristics of neural stem cells derived from adipose-derived stromal cells during their differentiation into neurons and astrocytes

AIM: To observe the ultrastructural characteristics of neural stem cells derived from adipose-derived stromal cells (ADSC) differentiating into neurons and astrocytes. METHODS: ADSC were cultured, amplified and induced with β-mercaptoethanol or 3-isobutyl-1-methylxanthine (IBMX). The expression of nestin in the induced cells was determined by the method of immunofluorescence. The ultrastructure of the induced neural stem cells was observed under transmission electron microscope.RESULTS: The expression of nestin reached the peak at 3 h in β-mercaptoethanol group and at 14 d in IBMX group, and the positive rate of the former (86%) was significantly higher than that of the latter (23%). However, the duration of the induction in IBMX group was obviously longer than that in β-mercaptoethanol group. The ultrastructure of the induced neural stem cells in the two groups was similar under the transmission electron microscope, only with some differences in organelles.CONCLUSION: In the process of ADSC differentiating into astrocytes or neurons with the induction of β-mercaptoethanol or IBMX, the different changes of ultrastructure in ADSC-derived neural stem cells occur in some organelles in cytoplasm, not in nucleus.     

Generation of insulin-producing cells from PDX1 and NKX6.1 gene-modified bone marrow mesenchymal stem cells

AIM: To differentiate bone marrow mesenchymal stem cells (BMSCs) into functional insulin-producing cells to produce sufficient pancreatic islet cells for transplantation. METHODS: Recombinant adenovirus vectors carrying PDX1 and NKX6.1 genes were constructed and the bone marrow mesenchymal stem cells were infected by the recombinant adenovirus combined with several cytokines for differentiation. The expressions of PDX1, NKX6.1 and insulin and C-peptide in the differentiated bone marrow mesenchymal stem cells were detected by RT-PCR and Western blotting. After the differentiated bone marrow mesenchymal stem cells were transplanted into subrenal capsule of diabetic mice, cell morphology of the grafts as well as their secretion of insulin and C-peptide were detected. Besides, regulating capacities of grafts on the blood glucose level of the diabetic mice were also detected. RESULTS: BMSCs induced by recombinant adenovirus (pAdxsi-CMV-PDX1/CMV-NKX6.1) and several cytokines showed positive dithizone staining and the expressions of β-cell related molecule such as insulin and glucose transporter 2 were detected by RT-PCR, which showed a sustaining and stable expression. The similar results were showed by Western blotting, immunohistochemical staining and indirect immunofluorescence. The insulin secretions in the cells stimulated with glucose at concentrations of 5.5 and 25 mmol/L in the experimental group were (1 240.4±109.3) mU/L and (3 539.8±245.1) mU/L, respectively, and were significantly higher than those in control group. Moreover, transplantation of the cells to STZ mice in treatment group made serum glucose recover to normal level. CONCLUSION: PDX1 and NKX6.1 gene-modified bone marrow mesenchymal stem cells differentiate into insulin-producing cells in vitro. When these cells transplanted into STZ induced diabetic mice, their serum glucose could return to the normal level and they could live well. Thus this is a promising method for diabetes treatment.