Electroacupuncture improves myocardial fibrosis in heart failure rats by attenuating ECM collagen deposition through modulation of TGF-β1/Smads signaling pathway

Background: To explore the effects of electroacupuncture on cardiac function and myocardial fibrosis in rat models of heart failure, and to elucidate the underlying mechanism of electroacupuncture in heart failure treatment. Methods: Healthy male Sprague-Dawley rats were allocated into three groups: Sham group, Model group, and electroacupuncture (Model + EA) group, with each group comprising 8 rats. The model underwent a procedure involving the ligation of the left anterior descending coronary artery to induce a model of heart failure. The Model + EA group was used for 7 consecutive days for electroacupuncture of bilateral Shenmen (HT7) and Tongli (HT5), once a day for 30 min each time. Left ventricular parameters in rats were assessed using a small-animal ultrasound machine to analyze changes in left ventricular end-diastolic volume, left ventricular end-systolic volume, left ventricular ejection fraction, and left ventricular fractional shortening. Serum interleukin-1β (IL-1β), cardiac troponin (cTn), and N-terminal brain natriuretic peptide precursor levels were measured using ELISA. Histopathological changes in rat myocardium were observed through HE staining, while collagen deposition in rat myocardial tissue was assessed using the Masson staining method. Picro sirius red staining, immunohistochemical staining, and RT-qPCR were utilized to distinguish between the various types of collagen deposition. The expression level of TGF-β1 and SMAD2/3/4/7 mRNA in rat myocardial tissues was determined using RT-qPCR. Additionally, western blot analysis was conducted to assess the protein expression levels of TGF-β1, SMAD3/7, and p-SMAD3 in rat myocardial tissues. Results: Compared with the Sham group, the left ventricular ejection fraction and left ventricular fractional shortening values of the Model group were significantly decreased (P < 0.01);the left ventricular end-diastolic volume and left ventricular end-systolic volume values were remarkably increased (P < 0.01);serum N-terminal brain natriuretic peptide precursor content was increased (P < 0.01);serum IL-1β and cTn levels were increased (P < 0.01);myocardial collagen volume fraction were increased (P < 0.01);and those of the expression of TGF-β1 and SMAD2/3/4 mRNA was increased (P < 0.01);the expression of SMAD7 mRNA was decreased (P < 0.01);the protein expression levels of TGF-β1, SMAD3, and p-Smad3 were increased (P < 0.01);the protein expression level of SMAD7 was decreased (P < 0.01) in the Model group. Compared to the Model group, the expression levels of the proteins TGF-β1, SMAD3, and p-Smad3 in myocardial tissue were found to be decreased (P < 0.01), and the expression level of the protein SMAD7 was found to be increased (P < 0.01) in the Model + EA group;the collagen volume fraction and deposition of type Ⅰ /Ⅲ collagen were decreased (P < 0.01) in the Model + EA group. Conclusion: Electroacupuncture alleviates myocardial fibrosis in rats with heart failure, and this effect is likely due to attributed to the modulation of the TGF-β1/Smads signaling pathway, which helps reduce collagen deposition in the extracellular matrix.

Interaction of major genes predisposing to hepatocellular carcinoma with genes encoding signal transduction pathways influences tumor phenotype and prognosis

Studies on rodents and humans demonstrate an inherited predisposition to hepatocellular carcinoma （HCC）. Analysis of the molecular alterations involved in the acquisition of a phenotype resistant or susceptible to hepatocarcinogenesis showed a deregulation of G1 and S phases in HCC of genetically susceptible F344 rats and a G1-S block in lesions of resistant Brown norway （BN） rats. Unrestrained extracellular signal-regulated kinase （ERK） activity linked to proteasomal degradation of dual-specificity phosphatase 1 （DUSP1）, a specific ERK inhibitor, by the CKS1-SKP2 ubiquitin ligase complex occurs in more aggressive HCC of F344 rats and humans. This mechanism is less active in HCC of BN rats and human HCC with better prognosis. Upregulation of iNos cross-talk with IKK/NF-KB and RAS/ERK pathways occurs in rodent liver lesions at higher levels in the most aggressive models represented by HCC of F344 rats and c-Myc-TGF-α transgenic mice. iNOS, IKK/NF-κB, and RAS/ERK upregulation is highest in human HCC with a poorer prognosis and positively correlates with tumor proliferation, genomic instability and microvascularization, and negatively with apoptosis. Thus, cell cycle regulation and the activity of signal transduction pathways seem to be modulated by HCC modifier genes, and differences in their efficiency influence the susceptibility to hepatocarcinogenesis and probably the prognosis of human HCC.

Sericin can reduce hippocampal neuronal apoptosis by activating the Akt signal transduction pathway in a rat model of diabetes mellitus

In the present study, a rat model of type 2 diabetes mellitus was established by continuous peritoneal injection of streptozotocin. Following intragastric perfusion of sericin for 35 days, blood glucose levels significantly reduced, neuronal apoptosis in the hippocampal CA1 region decreased, hippocampal phosphorylated Akt and nuclear factor kappa B expression were enhanced, but Bcl-xL/Bcl-2 associated death promoter expression decreased. Results demonstrated that sericin can reduce hippocampal neuronal apoptosis in a rat model of diabetes mellitus by regulating abnormal changes in the Akt signal transduction pathway.

Rho/Rock signal transduction pathway is required for MSC tenogenic differentiation

Mesenchymal stem cell （MSC）-based treatments have shown promise for improving tendon healing and repair. MSCs have the potential to differentiate into multiple lineages in response to select chemical and physical stimuli, including into tenocytes. Cell elongation and cytoskeletal tension have been shown to be instrumental to the process of MSC differentiation. Previous studies have shown that inhibition of stress fiber formation leads MSCs to default toward an adipogenic lineage, which suggests that stress fibers are required for MSCs to sense the environmental factors that can induce differentiation into tenocytes. As the Rho/ROCK signal transduction pathway plays a critical role in both stress fiber formation and in cell sensation, we examined whether the activation of this pathway was required when inducing MSC tendon differentiation using rope-like silk scaffolds. To accomplish this, we employed a loss-of-function approach by knocking out ROCK, actin and myosin （two other components of the pathway） using the specific inhibitors Y-27632, Latrunculin A and blebbistatin, respectively. We demonstrated that independently disrupting the cytoskeleton and the Rho/ ROCK pathway abolished the expression of tendon differentiation markers and led to a loss of spindle morphology. Together, these studies suggest that the tension that is generated by MSC elongation is essential for MSC teno-differentiation and that the Rho/ROCK pathway is a critical mediator of tendon differentiation on rope-like silk scaffolds.

Role of transforming growth factor-beta1-smad signal transduction pathway in patients with hepatocellular carcinoma

AIM： To explore the role of transforming growth factorbeta1 （TGF-β1）-smad signal transduction pathway in patients with hepatocellular carcinoma. METHODS： Thirty-six hepatocellular carcinoma specimens were obtained from Qidong Liver Cancer Institute and Department of Pathology of the Second Affiliated Hospital of Nanjing Medical University. All primary antibodies （polyclonal antibodies） to TGF-β1, type H Transforming growth factor-beta receptor （TβR-Ⅱ）, nuclear factor-kappaB （NF-KB）, CD34, smad4 and smad7, secondary antibodies and immunohistochemical kit were purchased from Zhongshan Biotechnology Limited Company （Beijing, China）. The expressions of TGF-β1, TβR-Ⅱ, NF- KB, smad4 and smad7 proteins in 36 specimens of hepatocellular carcinoma （HCC） and its adjacent tissue were separately detected by immunohistochemistry to observe the relationship between TGF-β1 and TβR-Ⅱ, between NF-KB and TGF-β1, between smad4 and smad7 and between TGF-β1 or TβR-Ⅱ and microvessel density （MVD）. MVD was determined by labelling the vessel endothelial cells with CD34. RESULTS： The expression of TGF-β1, smad7 and MVD was higher in HCC tissue than in adjacent HCC tissue （P〈0.01, P〈0.05,P〈0.01 respectively）. The expression of TβR-Ⅱ and smad4 was lower in HCC tissue than in its adjacent tissue （P〈0.01, P〈0.05 respectively）. The expression of TGF-β1 protein and NF-KB protein was consistent in HCC tissue. The expression of TGF-β1 and MVD was also consistent in HCC tissue. The expression of TIER- Ⅱ was negatively correlated with that of MVD in HCC tissue. CONCLUSION： The expressions of TGF-IB1, TβR- Ⅱ, NF-KB, smad4 and smad7 in HCC tissue, which are major up and down stream factors of TGF-β1-smad signal transduction pathway, are abnormal. These factors are closely related with NVD and may play an important role in HCC angiogenesis. The inhibitory action of TGF-β1 is weakened in hepatic carcinoma cells because of abnormality of TGF-β1 receptors （such as TIBR- Ⅱ） and postreceptors （such as smad4 and smad7）. NF-KB may cause activation and production of TGF-β1.

Role of Toll-like receptor 4 and Janus kinase and signal transducer and activator of transcription signal transduction pathway in sepsis-induced brain damage

The Janus kinase and signal transducer and activator of transcription （JAK/STAT） signal transduction pathway is involved in sepsis-induced functional damage to the heart, liver, kidney, and other organs. However, the cellular and molecular mechanisms underlying sepsis-induced brain damage remain elusive. In the present study, we found severe loss of neurons in the hippocampal CA1 region in rats with sepsis-induced brain damage following intraperitoneal injection of endotoxin, The expression of toll-like receptor 4, tumor necrosis factor a, and interleukin-6 was significantly increased in brain tissues following lipopolysaccharide exposure. AG490 （JAK2 antagonist） and rapamycin （STAT3 antagonist） significantly reduced neuronal loss and suppressed the increased expression of toll-like receptor 4, tumor necrosis factor a, and interleukin-6 in the hippocampal CA1 region in sepsis-induced brain damaged rats. Overall, these data suggest that blockade of the JAK/STAT signal transduction pathway is neuroprotective in sepsis-induced brain damage via the inhibition of toll-like receptor 4, tumor necrosis factor a, and interleukin-6 exoression.

Involvement of M3 Cholinergic Receptor Signal Transduction Pathway in Regulation of the Expression of Chemokine MOB-1, MCP-1 Genes in Pancreatic Acinar Cells

Whether M3 cholinergic receptor signal transduction pathway is involved in regulation of the activation of NF-κB and the expression of chemokine MOB-1, MCP-1genes in pancreatic acinar cells was investigated. Rat pancreatic acinar cells were isolated, cultured and treated with carbachol, atropine and PDTC in vitro. The MOB-1 and MCP-1 mRNA expression was detected by using RT-PCR. The activation of NF-κB was monitored by using electrophoretic mobility shift assay. The results showed that as compared with control group, M3 cholinergic receptor agonist (10 -3 mol/L, 10 -4 mol/L carbachol) could induce a concentration-dependent and time-dependent increase in the expression of MOB-1, MCP-1 mRNA in pancreatic acinar cells. After treatment with 10 -3 mol/L carbachol for 2 h, the expression of MOB-1, MCP-1 mRNA was strongest. The activity of NF-κB in pancreatic acinar cells was significantly increased (P<0.01) after treated with M3 cholinergic receptor agonist (10 -3 mol/L carbachol) in vitro for 30 min. Either M3 cholinergic receptor antagonist (10 -5 mol/L atropine) or NF-κB inhibitor (10 -2 mol/L PDTC) could obviously inhibit the activation of NF-κB and the chemokine MOB-1, MCP-1 mRNA expression induced by carbachol (P<0.05). This inhibitory effect was significantly increased by atropine plus PDTC (P<0.01). The results of these studies indicated that M3 cholinergic receptor signal transduction pathway was likely involved in regulation of the expression of chemokine MOB-1 and MCP-1genes in pancreatic acinar cells in vitro through the activation of NF-κB.

The Antidepressant Mechanism of JiaWeiWenDan Decoction Regulating p38MAPK-ERK5 Signal Transduction Pathway

Objective: To investigate the anti-depression mechanism of JiaWeiWenDan Decoction in regulating p38MAPK-ERK5 signal transduction pathway. Methods: Depression model rats were randomly divided into Blank Control Group, Model Control Group, Chinese Medicine Treatment Group, and Western Medicine Treatment Group (hereinafter referred to as Blank Group, Model Group, Chinese Medicine Group, and Western Medicine Group), with 48 rats in each group. The mice were treated with p38MAPK-ERK5 on the 7th day, 14th day and 21st day, respectively, and the mice were treated for 28 days. The key targets and cytokines in p38MAPK-ERK5 signal transduction pathway were detected. Results: Compared with the Blank Group, the expression of p38MAPKmRNA in the hippocampus of the Model Group was increased. The Chinese Medicine Group and Western Medicine Group could reduce the expression of p38MAPK mRNA (P P P P Conclusion: The anti-inflammatory effect of JiaWeiWenDan Decoction may be related to the regulation of p38MAPK-ERK5 signaling pathway. With the advance of the treatment week, the best effect was obtained when the treatment was started on the 7th day of modeling.

Signaling cross-talk between TGF-β/BMP and other pathways

Transforming growth factor-beta （TGF-β）/bone morphogenic protein （BMP） signaling is involved in the vast majority of cellular processes and is fundamentally important during the entire life of all metazoans. Deregulation of TGF-β/ BMP activity almost invariably leads to developmental defects and/or diseases, including cancer. The proper functioning of the TGF-β/BMP pathway depends on its constitutive and extensive communication with other signaling pathways, leading to synergistic or antagonistic effects and eventually desirable biological outcomes. The nature of such signaling cross-talk is overwhelmingly complex and highly context-dependent. Here we review the different modes of cross-talk between TGF-β/BMP and the signaling pathways of Mitogen-activated protein kinase, phosphatidylinositol-3 kinase/ Akt, Wnt, Hedgehog, Notch, and the interleukin/interferon-gamma/tumor necrosis factor-alpha cytokines, with an emphasis on the underlying molecular mechanisms.

Xinfuli Granule improves post-myocardial infarction ventricular remodelingand myocardial fibrosis in rats by regulating TGF-β/Smads signaling pathway

Recent clinical and experimental studies have confirmed the effects of Xinfuli Granule (XG), a compound Chinese medicine in the prevention and treatment of heart failure (HF). This study aimed to investigate the effects and the mechanisms of XG on ventricular reconstruction in rats with acute myocardial infarction (AMI).MethodsSprague-Dawley rats were subjected to left anterior descending branch ligation. The rats that survived 24 h were randomly assigned to five groups: medium-dose of XG group (MI+XGM), high-dose of XG group (MI+XGH), carvedilol group (MI+C), medium-dose of XG + carvedilol group (MI+C+XGM). Fourteen rats underwent identical surgical procedures without artery ligation, serving as sham controls. At 28 days, left ventricular weight to body weight (LVW/BW) and heart weight to body weight (HW/BW) were calculated; left ventricular ejection fraction (LVEF), left ventricular shortening fraction (LVFS), left ventricular internal diameter at systole (LVIDS) were measured by ultrasound; HE staining, Masson staining, and Sirius red staining were used to assess the myocardial pathological and physiological changes as well as myocardial fibrosis area and non-infarct zone I/III collagen ratio. Expression of Smad3 were detected and analyzed by Western blot, immunohistochemistry and immunofluorescence. P-Smad3, Smad2 and Smad7 in the TGF-β/Smads signaling pathway were also analyzed by Western blot.ResultsThe LVIDS (P < 0.01), HW/BW (P < 0.05), type I/III collagen ratio (P < 0.01) and myocardial collagen (P < 0.01) decreased significantly while the LVW/BW, LVFS (P < 0.05) increased significantly in MI+XGM group as compared with those in other groups. The expression of key signal molecules of the TGF-β/Smads signaling pathway, including Smad3, P-Smad3 and Smad2 protein were decreased, while the expression of Smad7 increased in both XG and carvedilol treatment groups as compared to those of the MI group (all P < 0.01). Immunohistochemistry and immunofluorescence further confirmed the down-regulated Smad3 expression.ConclusionXG can improve ventricular reconstruction and inhibit myocardial fibrosis in rats with AMI by regulating TGF-β/Smads signaling pathway.

MiR-146a-5p targeting SMAD4 and TRAF6 inhibits adipogenensis through TGF-β and AKT/mTORC1 signal pathways in porcine intramuscular preadipocytes

Background: Intramuscular fat(IMF) content is a vital parameter for assessing pork quality. Increasing evidence has shown that microRNAs(miRNAs) play an important role in regulating porcine IMF deposition. Here, a novel miRNA implicated in porcine IMF adipogenesis was found, and its effect and regulatory mechanism were further explored with respect to intramuscular preadipocyte proliferation and differentiation.Results: By porcine adipose tissue miRNA sequencing analysis, we found that miR-146a-5p is a potential regulator of porcine IMF adipogenesis. Further studies showed that miR-146a-5p mimics inhibited porcine intramuscular preadipocyte proliferation and differentiation, while the miR-146a-5p inhibitor promoted cell proliferation and adipogenic differentiation. Mechanistically, miR-146a-5p suppressed cell proliferation by directly targeting SMAD family member 4(SMAD4) to attenuate TGF-β signaling. Moreover, miR-146a-5p inhibited the differentiation of intramuscular preadipocytes by targeting TNF receptor-associated factor 6(TRAF6) to weaken the AKT/mTORC1 signaling downstream of the TRAF6 pathway.Conclusions: MiR-146a-5p targets SMAD4 and TRAF6 to inhibit porcine intramuscular adipogenesis by attenuating TGF-β and AKT/mTORC1 signaling, respectively. These findings provide a novel miRNA biomarker for regulating intramuscular adipogenesis to promote pork quality.

Berberine Attenuates Cigarette Smoke Extract-induced Airway Inflammation in Mice:Involvement of TGF-β1/Smads Signaling Pathway

Although several studies confirmed that berberine may attenuate airway inflammation in mice with chronic obstructive pulmonary disease(COPD),its underlying mechanisms were not clear until now.We aimed to establish an experiment mouse model for COPD and to investigate the effects of berberine on airway inflammation and its possible mechanism in COPD model mice induced by cigarette smoke extract(CSE).Twenty SPF C57BL/6 mice were randomly divided into PBS control group,COPD model group,low-dose berberine group and high-dose berberine group,5 mice in each group.The neutrophils and macrophages were examined by Wright's staining.The levels of inflammatory cytokines TNF-α and IL-6 in bronchoalveolar lavage fluid(BALF)were detennined by enzyme-linked immunosorbent assay.The expression levels of TGF-β1,Smad2 and Smad3 mRNA and proteins in lung tissues were respectively detected by quantitative real-time polymerase chain reaction and Western blotting.It was found that CSE increased the number of inflammation cells in BALF,elevated lung inflammation scores,and enhanced the TGF-β1/Smads signaling activity in mice.High-dose berberine restrained the alterations in the COPD mice induced by CSE.It was concluded that high-dose berberine ameliorated CSE-induced airway inflammation in COPD mice.TGF-β1/Smads signaling pathway might be involved in the mechanism.These findings suggested a therapeutic potential of high-dose berberine on the CSE-induced airway inflammation.

TGF-β1/SMAD SIGNALING PATHWAY MEDIATES p53-DEPENDENT APOPTOSIS IN HEPATOMA CELL LINES

Objective To determine whether transforming growth factor betal (TGF-β1)/Smad signaling pathway mediates p53-dependent apoptosis in hepatoma cell lines.Methods Three human hepatic carcinoma cell lines, HepG2, Huh-7, and Hep3B, were used in this study.TGF-β1-induced apoptosis in hepatic carcinoma cell lines was analyzed using TUNEL assay.For identifying the mechanism of apoptosis induced by TGF-β1, cell lines were transfected with a TGF-β1-inducible luciferase reportor plasmid containing Smad4 binding elements.After transfection, cells were treated with TGF-β1, then assayed for luciferase activity.Results The apoptosis rate of HepG2 cell lines (48.51%± 8.21%) was significantly higher than control ( 12.72%±2.18%, P<0.05).But TGF-β1 was not able to induce apoptosis of Huh-7 and Hep3B cell lines.The relative luciferase activity of TGF-β1-treated HepG2 cell lines (4.38) was significantly higher than control (1.00, P< 0.05).But the relative luciferase activity of TGF-β1-treated Huh-7 and Hep3B cell lines less increased compared with control.Conclusions HepG2 cells seem to be highly susceptible to TGF-β1-induced apoptosis compared with Hep3B and Huh-7 cell lines.Smad4 is a central mediator of TGF-β1 signaling transdution pathway.TGF-β1/Smad signaling pathway might mediate p53-dependent apoptosis in hepatoma cell lines.

Influence of electroacupuncture on mitogen-activated protein kinase signal transduction in a rat model of cerebral ischemia/reperfusion

Following electroacupuncture at Baihui （DU 20） and Dazhui （DU 14） in a rat model of cerebral ischemia/reperfusion, extracellular-signal-regulated kinase expression in cerebral cortex and corpus striatum, serum glutathione reductase, glutathione peroxidase activity, and serum glutathione content were elevated, and neurobehavioral scores improved. However, these effects were antagonized by mitogen-activated protein kinase inhibitor PD98059. Results indicated that electroacupuncture reversed free radical chain reactions and oxidative stress injury caused by cerebral ischemia/reperfusion, thereby providing neuroprotection. This process could correlate with the mitogen-activated protein kinase signal transduction pathway.

Cetirizine regulates scleroderma skin fibrosis in mice via the TGF-β1/Smad3 signaling pathway

Objective:To investigate the effect of cetirizine on the fibrosis of skin tissue in systemic sclerosis(SSc)mice and its mechanism of action.Methods:Thirty-two BALB/C mice were randomly divided into a blank group,a model group,a cetirizine low-dose group,and a cetirizine high-dose group,with eight in each group.The blank group was injected with normal saline on the back,and the other three groups were injected with bleomycin on the back to prepare SSc mouse models.The mice were injected once a day for 28 consecutive days,while the normal group and the model group were given saline.The dose group was administrated intragastrically at 2 mg/kg and 5 mg/kg,respectively,for 28 consecutive days.Detect the thickness of the dermis by taking the skin tissue in the back injection area of each group.Hematoxylin-eosin staining(HE)and Masson staining.Sample hydrolysis method to detect hydroxyproline(HYP)content in skin tissue.Immunohistochemical detection ofα-smooth muscle actin(α-SMA)expression in skin tissues.Enzyme-linked immunosorbent assay(ELISA)to detect serum interleukin(IL-6,IL-10)and transforming growth factor(TGF-αand TGF-β1).Quantitative real-time PCR(qRT-PCR)was used to detect the expression levels of collagen type I(COL1A1),type III collagen(COL3A1),Smad homolog 3(Smad3),and TGF-β1 mRNA.Western blot was used to detect the expression levels of COL1A1,COL3A1 and p-Smad3.Results:Compared with the blank group,the dermis thickness and HYP content of the model group increased,the skin tissue lesions and fibrosis were more severe,theα-SMA positive expression intensity in the skin tissue was higher,and the serum IL-6,IL-10,TGF-α,TGF-β1 content increased,COL1A1,COL3A1,Smad3,TGF-β1 mRNA expression levels increased in skin tissues,COL1A1,COL3A1,p-Smad3 protein expression increased,the differences were statistically significant(P<0.05).Compared with the model group,the dermal thickness and HYP content of the low and high dose cetirizine groups were reduced,the degree of skin tissue lesions and fibrosis was improved,the expression ofα-SMA in skin tissues was weakened,the levels of IL-6,IL-10,TGF-α,TGF-β1 in serum were reduced,the expression levels of COL1A1,COL3A1,Smad3 and TGF-β1 in skin tissues were reduced,and the expression levels of COL1A1,COL3A1,and p-Smad3 proteins were reduced,the decrease in the high-dose group was more significant,and the differences were statistically significant(P<0.05).Conclusion:Cetirizine can improve the degree of fibrosis of skin tissue in SSc mice and reduce the immune inflammation response.The mechanism of action is related to the TGF-β1/Smad3 signaling pathway.

Effects of Cigu Xiaozhi Formula on miR-378a-3p Expression and Hh Signaling Pathway in TGF-β1 Induced LX2 Cells

[Objectives]To observe the effects of Cigu Xiaozhi Formula on miR-378a-3p expression and Hh signaling pathway in TGF-β1 induced and activated LX2 cells.[Methods]Cells were divided into control group,induction group,drug-containing serum group,miR-378a-3p inhibitor group,and miR inhibitor NC group.CCK-8 method was used to detect the cell viability of each group,and flow cytometry was used to detect the apoptosis rate of each group.RT-qPCR was used to detect the expression of miR-378a-3p in each group s cells,and RT-qPCR and Western blot were used to detect mRNA and protein expression of Shh,Gli1,Gli2,Col-I,andα-SMA in each group s cells.[Results]Compared with the control group,the cell viability and expression of Shh,Gli1,Gli2,Col-I,andα-SMA mRNA and protein in induction group increased(P<0.01),while the expression of miR-378a-3p decreased(P<0.01).Compared with the induction group,the cell viability and expression of Shh,Gli1,Gli2,Col-I,α-SMA mRNA andα-SMA and Gli2 protein decreased in drug-containing serum group(P<0.05),while cell apoptosis rate and miR-378a-3p expression increased(P<0.01).In miR-378a-3p inhibitor group,cell viability and the expression of Shh,Gli1,Gli2,Col-I,α-SMA mRNA and Gli1,Gli2,α-SMA protein increased(P<0.05,P<0.01),while the apoptosis rate and miR-378a-3p expression decreased(P<0.05,P<0.01).[Conclusions]Cigu Xiaozhi Formula containing serum can upregulate miR-378a-3p expression and downregulate the expression of Gli2 andα-SMA in TGF-β1 induced LX2 cells,thereby inhibiting the activation of LX2 cells and exerting the effects of anti liver fibrosis.

Ski and SnoN, potent negative regulators of TGF-β signaling

Ski and the closely related SnoN were discovered as oncogenes by their ability to transform chicken embryo fibroblasts upon overexpression. While elevated expressions of Ski and SnoN have also been reported in many human cancer cells and tissues, consistent with their pro-oncogenic activity, emerging evidence also suggests a potential anti-oncogenic activity for both. In addition, Ski and SnoN have been implicated in regulation of cell differentiation, especially in the muscle and neuronal lineages. Multiple cellular partners of Ski and SnoN have been identifed in an effort to understand the molecular mechanisms underlying the complex roles of Ski and SnoN. In this review, we summarize recent findings on the biological functions of Ski and SnoN, their mechanisms of action and how their levels of expression are regulated.

Role of TGF-βl Signaling in Heart Valve Calcification Induced by Abnormal Mechanical Stimulation in a Tissue Engineering Model

A tissue engineering model of heart valve calcification induced in a bio-reactor was established to evaluate the calcification induced by abnormal mechanical stimulation and explore the underlying molecular mechanisms.Polyethylene glycol (PEG)-modified decellularized porcine aortic leaflets seeded with human valve interstitial cells (huVICs)were mounted on a Ti-Ni alloy frame to fabricate two-leaflet and three-leaflet tissue engineered valves.The two-leaflet model valves were exposed to abnormal pulsatile flow stimulation with null (group A),low (1000mL/min,group B),medium (2000mL/min,group C),and high velocity (3000mL/min,group D)for 14 days. Morphology and calcification were assessed by yon Kossa staining,alkaline phosphatase (ALP)content,and Runx2 immunostaining.Leaflet calcification and mRNA and protein expression of transforming growth factor (TGF)-β1,bone morphogenetic protein 2 (BMP2),Smadl,and MSX2 were measured at different time points.ALP content was examined in two-leaflet valves seeded with BMP2 shRNA plasmid-infected huVICs and exposed to the same stimulation conditions.The results showed that during 14 days of flow stimulation,huVICs on the leaflet surface proliferated to generate normal monolayer coverage in groups A,B,and C.Under mechanical stimulation,huVICs showed a parallel growth pattern in the direction of the fluid flow,but huVICs exhibited disordered growth in the high-velocity flow environment,yon Kossa staining,ALP measurement,and immunohistochemical staining for Runx2 confirmed the lack of obvious calcification in group A and significant calcification in group D.Expression levels of TGF-β1,BMP2, and MSX2 mRNA and protein were increased under fluid stimulation.ALP production by BMP2 shRNA plasmid-infected huVICs on model leaflets was significantly reduced.In conclusion,abnormal mechanical stimulation in a bioreactor induced calcification in the tissue engineering valve model.The extent of calcification correlated positively with the flow velocity,as did the mRNA and protein levels of TGF-β1,BMP2,and MSX2.These findings indicate that TGF-β1/BMP2 signaling is involved in valve calcification induced bv abnormal mechanical stimulation.

Mitogen activated protein kinase signaling pathways participate in the active principle region of Buyang Huanwu decoction-induced differentiation of bone marrow mesenchymal stem cells

Our preliminary studies confirmed that an active principle region of Buyang Huanwu decoction, comprising alkaloid, polysaccharide, aglycon, glucoside and volatile oil, can induce bone marrow mesenchymal stem cell differentiation into neurons. Mitogen-activated protein kinase signaling was identified as one of the key pathways underlying this differentiation process. The present study shows phosphorylated extracellular signal-regulated protein kinase and phosphorylated p38 protein expression was increased after differentiation. Cellular signaling pathway blocking agents, PD98059 and SB203580, inhibited extracellular signal-regulated protein kinase and p38 in mitogen-activated protein kinase signaling pathways respectively, mRNA and protein expression of the neuronal marker, neuron specific enolase, and neural stem cell marker, nestin, were decreased in bone marrow mesenchymal stem cells after treatment with the active principle region of Buyang Huanwu decoction. Experimental findings indicate that, extracellular signal-regulated protein kinase and p38 in mitogen-activated protein kinase signaling pathways participate in bone marrow mesenchymal stem cell differentiation into neuron-like cells, induced by the active principle region of Buyang Huanwu decoction.