Transforming growth factor-β1 and vascular endothelial growth factor levels in senile acute myeloid leukemia and correlation with prognosis

BACKGROUND Acute myeloid leukemia(AML)is a disease in which immature hematopoietic cells accumulate in the bone marrow and continuously expand,inhibiting hematopoiesis.The treatment and prognosis of this disease have always been unsatisfactory.AIM To investigate the correlation between vascular endothelial growth factor(VEGF)and transforming growth factor-β1(TGFβ1)expression and prognosis in older adults with AML.METHODS This study enrolled 80 patients with AML(AML group),including 36 with complete response(AML-CR),23 with partial response(AML-PR),and 21 with no response(AML-NR).The expression levels of VEGF and TGFβ1 were detected by reverse transcription polymerase chain reaction in bone marrow mononuclear cells isolated from 56 healthy controls.Kaplan-Meier analysis was performed to assess overall survival(OS)and progression-or disease-free survival(DFS).Prognostic risk factors were analyzed using a Cox proportional hazards model.RESULTS The AML group showed a VEGF level of 2.68±0.16.VEGF expression was lower in patients with AML-CR than those with AML-PR or AML-NR(P<0.05).TGFβ1 expression in the AML group was 0.33±0.05.Patients with AML-CR showed a higher TGFβ1 expression than those with AML-PR or AML-NR(P<0.05).VEGF and TGFβ1 expression in patients with AML was significantly correlated with the counts of leukocytes,platelets,hemoglobin,and peripheral blood immature cells(P<0.05);Kaplan-Meier survival analysis revealed that patients with high TGFβ1 expression had better OS and DFS than those with low TGFβ1 expression(P<0.05),whereas patients with low VEGF levels showed better OS and DFS than those with high VEGF levels(P<0.05).VEGF,TGFβ1,and platelet count were identified by the Cox proportional hazards model as independent risk factors for OS(P<0.05),while VEGF,TGFβ1,and white blood cell count were independent risk factors for DFS(P<0.05).CONCLUSION Decreased VEGF expression and increased TGFβ1 expression in patients with AML provide valuable references for determining and individualizing clinical treatment strategies.

Plasma Levels of Transforming Growth Factor-Beta 1 in Women with Pelvic Organ Prolapse

Objective: In women with pelvic organ prolapse (POP), decreased expression of transforming growth factor-beta 1 (TGF-β1) has been shown in POP tissues. However, no studies have evaluated plasma TGF-β1 levels in patients with POP, so it is unknown whether they are also changed or not. Therefore, we compared plasma TGF-β1 levels in women with and without POP. Methods: Participants were 49 women with POP and 23 healthy control women. All participants were postmenopausal. We measured plasma TGF-β1 and compared data between patients with POP and controls, and between patients with uterine prolapse (UP, n = 19) and those with a cystocele (CC, n = 30). In addition, in patients, we assessed the POP quantification system (POP-Q) stage. Results: Plasma TGF-β1 levels were significantly lower in patients than in healthy controls. POP-Q stage was not significantly different between the UP and CC subgroups, but POP-Q stage IV was diagnosed in 63% of patients with UP and 7% of those with CC. Plasma TGF-β1 levels were significantly lower in the CC subgroup than in the UP subgroup. Conclusion: Plasma TGF-β1 is decreased in POP. It remains unclear whether the lower levels indicate a reduction in systemic TGF-β1 activity, but they can be assumed to reflect reduced TGF-β1 expression in POP tissues.

Expression of transforming growth factor-β_1 and its typeⅠ receptor in different phases of post-burn hypertrophic scars

Objective: To analyze and compare the expression pattern of the transforming growth factor-β1(TGF-β1) and its type I receptor (TGF-β RI ) in nounal human skin and various phases of post-burn hypertrophic scars (HTS). Method: The immunohistochemical ABC method was employed. Results: In nounal human skin, no evident immunoreactivity of TGF-β1 and TGF-β R I was observed. In activation phase of post-burn HTS, TGF-β R I and TGF-β1 were highly expressed in most dermal fibroblasts which seemed to be the same subset. However, in remission phase, no staining was seen in der mal fibroblasts. Conclusion: The formation of all may involve the increase of TGF-β responsiveness in fibroblasts The ac cumulation at the wound site and failure of apoptosis of over-resposive fibroblasts may contribute to the formation of HTS.

Effects of Heparin on Transforming Growth Factor-β_1 and Extracellular Matrix Components in the Glomeruli of Diabetic Rats

The effects of heparin on the expression of transforming growth factor-β 1 (TGF-β 1) and two extracellular matrix components laminin (LN) and fibronectin (FN) in diabetic rat glomeruli were investigated. Twenty-six rats were randomly divided into control group (C, n=8), diabetic group (D, n=9), and diabetes+heparin group (DH, n=9). After 8-week therapy of heparin (200 U once daily by abdominal injection), TGF-β 1, LN and FN expression in glomeruli was detected by immunohistochemical method. The results showed that the expression levels of TGF-β 1, LN and FN were higher in group D than in group C. It was found that heparin could reduce 24-h urinary albumin excretion and inhibit overexpression of TGF-β 1, LN and FN in glomeruli of diabetic rats. It suggested that the inhibitory effect of heparin on diabetic glomerular sclerosis was at least partly related with the inhibition of TGF-β 1 expression.

Expression of Mesenger RNA for Transforming Growth Factor-β_1 in Bovine Trabecular Meshwork

Purpose: To investigate the relationship between transforming growth factor-β1(TGF-β1) and primary open-angle glaucoma, we have determined whether trabec-ular tissues have the expression of messenger RNA for TGF-β1.Methods: Total RNA of 24 newborn bovine trabecular tissue were extracted byGuanidine isothiocyanate method. The TGF-β33 plasmid was brought into E. col-ibacillius HB101 and amplificated. After Bam HI endolase degradation and labelwith a-32p-dATP the RNA was hybridized with the cDNA (complementary DNA)probe and examined by autoradiography.Results: The presence of mRNA for TGF-β1 in bovine trabecular meshwork wasconfirmed.Conclusions: The TGF-β1 present in normal aqueous humor must be at least partlyderived from the trabecular meshwork. It offered a basis for understanding therelationship between abnormal synthesis, activation and clearance of TGF-β1 andthe pathogenesis of primary open-angle glaucoma (POAG) in molecular biology.Eye Science 1996; 12:1-4.

TL1A Promotes Fibrogenesis in Colonic Fibroblasts via the TGF-β1/Smad3 Signaling Pathway

Objective Intestinal fibrosis is a refractory complication of inflammatory bowel disease(IBD).Tumor necrosis factor ligand-related molecule-1A(TL1A)is important for IBD-related intestinal fibrosis in a dextran sodium sulfate(DSS)-induced experimental colitis model.This study aimed to explore the effects of TL1A on human colonic fibroblasts.Methods A trinitrobenzene sulfonic acid(TNBS)-induced experimental colitis model of LCK-CD2-TL1A-GFP transgenic(Tg)or wild-type(WT)mice was established to determine the effect and mechanism of TL1A on intestinal fibrosis.The human colonic fibroblast CCD-18Co cell line was treated concurrently with TL1A and human peripheral blood mononuclear cell(PBMC)supernatant.The proliferation and activation of CCD-18Co cells were detected by BrdU assays,flow cytometry,immunocytochemistry and Western blotting.Collagen metabolism was tested by Western blotting and real-time quantitative polymerase chain reaction(RT-qPCR).Results The level of collagen metabolism in the TNBS+ethyl alcohol(EtOH)/Tg group was greater than that in the TNBS+EtOH/WT group.Transforming growth factor-β1(TGF-β1)and p-Smad3 in the TNBS+EtOH/Tg group were upregulated as compared with those in the TNBS+EtOH/WT group.The proliferation of CCD-18Co cells was promoted by the addition of human PBMC supernatant supplemented with 20 ng/mL TL1A,and the addition of human PBMC supernatant and TL1A increased CCD-18Co proliferation by 24.4%at 24 h.TL1A promoted cell activation and increased the levels of COL1A2,COL3A1,and TIMP-1 in CCD-18Co cells.Treatment of CCD-18Co cells with TL1A increased the expression of TGF-β1 and p-Smad3.Conclusion TL1A promotes TGF-β1-mediated intestinal fibroblast activation,proliferation,and collagen deposition and is likely related to an increase in the TGF-β1/Smad3 signaling pathway.

Unveiling the therapeutic potential:KBU2046 halts triple-negative breast cancer cell migration by constricting TGF-β1 activation in vitro

Background:Triple-negative breast cancer(TNBC)is a heterogeneous,recurring cancer characterized by a high rate of metastasis,poor prognosis,and lack of efficient therapies.KBU2046,a small molecule inhibitor,can inhibit cell motility in malignant tumors,including breast cancer.However,the specific targets and the corresponding mechanism of its function remain unclear.Methods:In this study,we employed(3-(4,5-dimethylthiazol-2-yl)-5-(3-carboxymethoxyphenyl)-2-(4-sulfophenyl)-2H tetrazolium)(MTS)assay and transwell assay to investigate the impact of KBU2046 on the proliferation and migration of TNBC cells in vitro.RNA-Seq was used to explore the targets of KBU2046 that inhibit the motility of TNBC.Finally,confirmed the predicted important signaling pathways through RT-qPCR and western blotting.Results:In this study,we found that KBU2046 functioned as a novel transforming growth factor-β(TGF-β1)inhibitor,effectively suppressing tumor cell motility in vitro.Mechanistically,it directly down-regulated leucine-rich repeat-containing 8 family,member E(LRRC8E),latent TGFβ-binding protein 3(LTBP3),dynein light chain 1(DNAL1),and MAF family of bZIP transcription factors(MAFF)genes,along with reduced protein expression of the integrin family.Additionally,KBU2046 decreased phosphorylation levels of Raf and ERK.This deactivation of the ERK signaling pathway impeded cancer invasion and metastasis.Conclusions:In summary,these findings advocate for the utilization of TGF-β1 as a diagnostic and prognostic biomarker and as a therapeutic target in TNBC.Furthermore,our data underscore the potential of KBU2046 as a novel therapeutic strategy for combating cancer metastasis.

Vascular endothelial growth factor A, secreted in response to transforming growth factor-β1 under hypoxic conditions, induces autocrine effects on migration of prostate cancer cells

Hypoxia and transforming growth factor-β1 （TGF-β1） increase vascular endothelial growth factor A （VEGFA） expression in a number of malignancies. This effect of hypoxia and TGF-β1 might be responsible for tumor progression and metastasis of advanced prostate cancer. In the present study, TGF-β1 was shown to induce VEGFA165 secretion from both normal cell lines （HPV7 and RWPE1） and prostate cancer cell lines （DU 145 and PC3）. Conversely, hypoxia-stimulated VEGFA165 secretion was observed only in prostate cancer cell lines. Hypoxia induced TGF-β1 expression in PC3 prostate cancer cells, and the TGF-β1 type I receptor （ALK5） kinase inhibitor partially blocked hypoxia-mediated VEGFA16s secretion. This effect of hypoxia provides a novel mechanism to increase VEGFA expression in prostate cancer cells. Although autocrine signaling of VEGFA has been implicated in prostate cancer progression and metastasis, the associated mechanism is poorly characterized. VEGFA activity is mediated via VEGF receptor （VEGFR） 1 （Fit-l） and 2 （FIk-I/KDR）. Whereas VEGFR-1 mRNA was detected in normal prostate epithelial cells, VEGFR-2 mRNA and VEGFR protein were expressed only in PC3 cells. VEGFA165 treatment induced phosphorylation of extracellular signal-regulated kinase 1/2 （ERKI/2） in PC3 cells but not in HPV7 cells, suggesting that the autocrine function of VEGFA may be uniquely associated with prostate cancer. Activation of VEGFR-2 by VEGFA165 was shown to enhance migration of PC3 cells. A similar effect was also observed with endogenous VEGFA induced by TGF-β1 and hypoxia. These findings illustrate that an autocrine loop of VEGFA via VEGFR-2 is critical for the tumorigenic effects of TGF-β1 and hypoxia on metastatic prostate cancers.

TRANSFORMING GROWTH FACTOR-β1 AND SMAD4 SIGNALING PATHWAY DOWN-REGULATES RENAL EXTRACELLULAR MATRIX DEGRADATION IN DIABETIC RATS

Objective To investigate the role of transforming growth factor-131 （TGF-β1）/Smad4 pathway in development of renal fibrosis in streptozotocin （STZ）-induced diabetic nephropathy （DN） rats and explore its possible mechanism. Methods Male Wistar rats weighing 180-220 g were divided into 5 groups： group A （ normal control）, group B [ diabetes mellitus （DM） 2 weeks ], group C （ DM 4 weeks）, group D （ DM 8 weeks）, and group E （ DM 16 weeks）. Except for the normal control group, other groups were induced DM by single injection of STZ （55 mg/kg） respectively. Blood glucose level, serum creatinine, and 24-hour urine protein were examined. Expressions of TGF-β1 and Smad4 protein and mRNA in kidney were detected using immunohistochemical technique, Western blot, and real-time PCR. mRNA expressions of stromelysin-1 （ MMP-3 ）, tissue inhibitor of metalloproteinase-1 （ TIMP-1 ）, and collagen Ⅲ in kidney were also detected by real-time PCR. Results The levels of blood glucose, serum creatinine, and 24-hour urine protein in rats of group B, C, D, and E were higher than those of the control group. With the progression of renal fibrosis, the expressions of TGF-β1 and Smad4 protein and mRNA in kidney of diabetic rats elevated. In addition, the renal MMP-3 mRNA expression diminished in diabetic rats, while TIMP-1 and collagen Ⅲ mRNA increased. Conclusions In STZ-induced diabetic rats, the TGF-β1/Smad4 appears to play an important role in renal fibrosis of DN. The increased expression of TGF-β1 and Smad4 might result in the transcriptional regulation of downstream target genes of TGF-β1/Smad4 pathway, which contributes to the progression of renal fibrosis in diabetic rats.

The Effect of Simvastatin on mRNA Expression of Transforming Growth Factor-β1,Bone Morphogenetic Protein-2 and Vascular Endothelial Growth Factor in Tooth Extraction Socket

Aim To determine the effect of local simvastatin application on the mRNA expression level of transforming growth factor-β1 （TGF-β1）, bone morphogenetic protein-2 （BMP-2） and vascular endothelial growth factor （VEGF） in the tooth sockets of rat. Methodology Forty-eight male Wistar rats were randomly divided into experimental and control groups （n=24）. Polylactic acid/polyglycolic acid copolymer carriers, with or without simvastatin, were implanted into extraction sockets of right mandibular incisors. The expression of TGF-β1, BMP-2 and VEGF mRNA was determined by in situ hybridization in the tooth extraction socket at five days, one week, two weeks and four weeks after implantation. Results The fusiform stroma cells in the tooth extraction socket began to express TGF-β1, BMP-2 and VEGF mRNA in both experimental and control groups from one week after tooth extraction until the end of experiment. The expression of TGF-131 and BMP-2 mRNA in the experimental group was significantly up-regulated after one, two and four weeks, and expression of VEGF mRNA was significantly increased after one and two weeks compared with that in the control group. Conclusion The findings indicate that local administration of simvastatin can influence alveolar bone remodeling by regulating the expression of a school of growth factors which are crucial to osteogenesis in the tooth extraction socket.

Total flavone of Abelmoschus manihot suppresses epithelial-mesenchymal transition via interfering transforming growth factor-β1 signaling in Crohn's disease intestinal fibrosis

AIM To explore the role and mechanism of total flavone of Abelmoschus manihot(TFA) on epithelial-mesenchymal transition(EMT) progress of Crohn's disease(CD) intestinal fibrosis.METHODS First,CCK-8 assay was performed to assess TFA on the viability of intestinal epithelial(IEC-6) cells and select the optimal concentrations of TFA for our further studies.Then cell morphology,wound healing and transwell assays were performed to examine the effect of TFA on morphology,migration and invasion of IEC-6 cells treated with TGF-β1.In addition,immunofluorescence,real-time PCR analysis(q RT-PCR) and western blotting assays were carried out to detect the impact of TFA on EMT progress.Moreover,western blotting assay was performed to evaluate the function of TFA on the Smad and MAPK signaling pathways.Further,the role of co-treatment of TFA and si-Smad or MAPK inhibitors has been examined by q RTPCR,western blotting,morphology,wound healing andtranswell assays.RESULTS In this study,TFA promoted transforming growth factor-β1(TGF-β1)-induced(IEC-6) morphological change,migration and invasion,and increased the expression of epithelial markers and reduced the levels of mesenchymal markers,along with the inactivation of Smad and MAPK signaling pathways.Moreover,we revealed that si-Smad and MAPK inhibitors effectively attenuated TGF-β1-induced EMT in IEC-6 cells.Importantly,co-treatment of TFA and si-Smad or MAPK inhibitors had better inhibitory effects on TGF-β1-induced EMT in IEC-6 cells than either one of them.CONCLUSION These findings could provide new insight into the molecular mechanisms of TFA on TGF-β1-induced EMT in IEC-6 cells and TFA is expected to advance as a new therapy to treat CD intestinal fibrosis.

Promoter polymorphism of transforming growth factor-β1 gene and ulcerative colitis

AIM: To elucidate the possible difference in two promoter polymorphisms of the transforming growth factor-β1 (TGF-β1) gene (-800G > A, -509C > T) between ulcerative colitis (UC) patients and normal subjects.METHODS: A total of 155 patients with established ulcerative colitis and 139 normal subjects were selected as controls. Two single nucleotide polymorphisms within the promoter region of TGF-β1 gene (-509C > T and -800G > A) were genotyped using PCR-RFLP. RESULTS: There was a statistically significant difference in genotype and allele frequency distributions between UC patients and controls for the -800G > A polymorphism of the TGF-β1 gene (P < 0.05). The frequency of the TGF-β1 gene polymorphism at position -800 showed that the AA genotype and the allele A frequencies significantly differed between the patients and healthy controls (P < 0.05). At position -509, there was no statically significant difference in genotype and allele frequency between the patients and control subjects.CONCLUSION: The results of our study indicate that there is a significant difference in both allele and genotype frequency at position -800G > A of TGF-β1 gene promoter between Iranian patients with UC and normal subjects.

Histone deacetylase inhibitor suberoylanilide hydroxamic acid alleviates liver fibrosis by suppressing the transforming growth factor-β1 signal pathway

Background: Histone deacetylases(HDACs) inhibitors are new anti-fibrotic drugs that inhibit the activity of hepatic stellate cells. The present study focused on the anti-fibrotic function of HDAC inhibitor suberoylanilide hydroxamic acid(SAHA) by suppressing transforming growth factor-β1(TGF-β1) signaling. Methods: Male Sprague-Dawley rats were used to induce liver fibrosis with carbon tetrachloride(CCl 4) and LX2 cell(human hepatic stellate cell line) was stimulated by TGF-β1. Both animals and cells were treated with SAHA. The Smad7 and connective tissue growth factor(CTGF) mRNA levels were detected by real-time polymerase chain reaction(PCR). Western blotting was used to examine the protein levels of CTGF, Histone H3(H3), Smad7, Smad2/3, Acetyl-Histone H3(AH3), HDAC2, α-smooth muscle actin( α-SMA), HDAC6, p-Smad2/3 and HDAC8. In addition, the TGF-β1 and liver enzyme levels from rat serum were detected. Histopathological changes were examined by hematoxylin and eosin(HE), Sirius red and Masson trichrome staining. The α-SMA expression was detected by immumohistochemical staining. Results: Compared with control group, the TGF-β1 and liver enzyme levels from rat serum, together with the mRNA levels of CTGF and protein levels of CTGF, HDAC2, α-SMA, HDAC6, p-Smad2/3 and HDAC8 were elevated in fibrotic rats( P < 0.01). But the Smad7 mRNA and AH3 protein levels were notably suppressed in the fibrotic rats( P < 0.01). Pathological examination showed the typical changes of liver fibrosis in the fibrotic rats. After the treatment with SAHA, the levels of liver enzymes, TGF-β1, CTGF, HDAC2, α-SMA, HDAC6, p-Smad2/3 and HDAC8 were reduced( P < 0.01) and Smad7 and AH3 protein contents were elevated in liver fibrotic rats( P < 0.01). Moreover, immumohistochemistry showed that SAHA significantly suppressed the α-SMA protein content in fibrotic liver( P < 0.01). Conclusion: The HDAC inhibitor SAHA alleviated liver fibrosis by suppressing the TGF-β1 signaling.

Dab2 attenuates brain injury in APP/PS1 mice via targeting transforming growth factor-beta/SMAD signaling

Transforming growth factor-beta （TGF-β） type II receptor （TβRⅡ） levels are extremely low in the brain tissue of patients with Alzheimer＇s disease. This receptor inhibits TGF-β1/SMAD signaling and thereby aggravates amyolid-beta deposition and neuronal injury. Dab2, a specific adapter protein, protects T RII from degradation and ensures the effective conduction of TGF-β 1/SMAD signaling. In this study, we used an adenoviral vector to overexpress the Dab2 gene in the mouse hippocampus and investigated the regulatory effect of Dab2 protein on TGF-β1/SMAD signaling in a mouse model of Alzheimer＇s disease, and the potential neuroprotective effect. The results showed that the TβRⅡ level was lower.in APP/PS1 mouse hippocampus than in normal mouse hippocampus. After Dab2 expression, hippocampal TβRⅡ and p-SMAD2/3 levels were signifi- cantly increased, while amyloid-beta deposition, microglia activation, tumor necrosis factor- and interleulin-6 levels and neuronal loss were significantly attenuated in APP/PS1 mouse brain tissue. These results suggest that Dab2 can exhibit neuroprotective effects in Alzheimer＇s disease by regulating TGF-β1/SMAD signaling.

Transactivation of the TIEG1 confers growth inhibition of transforming growth factor-β-susceptible hepatocellular carcinoma cells

AIM:To investigate the role of transforming growth factor(TGF)-β-inducible early gene 1(TIEG1) in TGF-β-induced growth inhibition in hepatocellular carcinoma(HCC) cells.METHODS:Human hepatocyte and HCC cell lines with varied susceptibilities to TGF-β1 were tested by methylthiazoletetrazolium(MTT) assay.The expression changes of Smad2,Smad3,Smad4,Smad7,TIEG1 and TIEG2 gene following treatment with TGF-β1 in a TGF-β-sensitive hepatocyte cell line(MIHA),a TGF-β-sensitive hepatoma cell line(Hep3B) and two TGF-β-insensitive hepatoma cell lines(HepG2 and Bel7404) were examined.SiRNA targeting TIEG1 was transfected into Hep3B cells and the sensitivity of cells to TGF-β1 was examined.Overexpression of TIEG1 was induced by lentiviral-mediated transduction in TGF-β1-resistant hepatoma cell lines(Bel7404 and HepG2).MTT assay and 4',6-Diamidino-2-phenylindole staining were used to identify cell viability and apoptosis,respectively.The expression level of stathmin was measured by reverse transcriptase polymerase chain reaction and Western-blotting analysis,and stathmin promoter activity by TIEG1 was monitored by a luciferase reporter gene system.RESULTS:TIEG1 was significantly upregulated by TGF-β1 in the TGF-β1-sensitive HCC cell line,Hep3B,but not in the resistant cell lines.The suppression of TIEG1 by siRNAs decreased the sensitivity of Hep3B cells to TGF-β1,whereas the overexpression of TIEG1 mediated growth inhibition and apoptosis in TGF-β1-resistant HCC cell lines,which resembled those of TGF-β1-sensitive HCC cells treated with TGF-β1.Our data further suggested that stathmin was a direct target of TIEG1,as stathmin was signif icantly downregulated by TIEG1 overexpression,and stathmin promoter activity was inhibited by TIEG1 in a dose-dependent manner.CONCLUSION:Our data suggest that transactivation of TIEG1 conferred growth inhibition of TGF-β-susceptible human HCC cells.

Effect of transforming growth factor-β1 on human intrahepatic cholangiocarcinoma cell growth

AIM： To elucidate the biological effects of transforming growth factor-β1 （TGF-β1） on intrahepatic cholangiocarcinoma （ICC）.METHODS： We investigated the effects of TGF-β1 on human ICC cell lines （HuCCT1, MEC, and HUH-28） by monitoring the influence of TGF-β1 on tumor growth and interleukin-6 （IL-6） expression in ICC cells.RESULTS： All three human ICC cell lines produced TGF-β1 and demonstrated accelerated growth in the presence of TGF-β1 with no apoptotic effect. Studies on HuCCT1 revealed a TGF-β1-induced stimulation of the expression of TGF-β1, as well as a decrease in TGF-β1 mRNA expression induced by neutralizing anti-TGF-β1 antibody. These results indicate that TGF-β1 stimulates the production and function of TGF-β1 in an autocrine fashion. Further, IL-6 secretion was observed in all three cell lines and exhibited an inhibitory response to neutralizing anti-TGF-β1 antibody. Experiments using HuCCT1 revealed a TGF-β1-induced acceleration of IL-6 protein expression and mRNA levels. These findings demonstrate a functional interaction between TGF-β1 and IL-6. All three cell lines proliferated in the presence of IL-6. In contrast, TGF-β1 induced no growth effect in HuCCT1 in the presence of small interfering RNA against a specific cell surface receptor of IL-6 and signal transducer and activator of transcription-3.CONCLUSION： ICC cells produce TGF-β1 and confer a TGF-β1-induced growth effect in an autocrine fashion.TGF-β1 activates ILo6 production, and the functional interaction between TGF-β1 and IL-6 contributes to ICC cell growth by TGF-β1.

Hsa_circRNA_102610 upregulation in Crohn’s disease promotes transforming growth factor-β1-induced epithelial-mesenchymal transition via sponging of hsa-miR-130a-3p

BACKGROUND The incidence of inflammatory bowel disease,a chronic intestinal inflammatory disorder that includes Crohn’s disease(CD)and ulcerative colitis,is rising.Circular RNAs are considered valuable diagnostic biomarkers for CD.Current evidence supports the views that epithelial-mesenchymal transition(EMT)plays an important role in CD pathogenesis,and that hsa-miR-130a-3p can inhibit transforming growth factor-β1(TGF-β1)-induced EMT.Our previous study revealed that hsa_circRNA_102610 was upregulated in CD patients.Moreover,we predicted an interaction between hsa_circRNA_102610 and hsa-miR-130a-3p.Thus,we hypothesized that hsa_circRNA_102610 may play roles in the proliferation and EMT of intestinal epithelial cells by sponging hsa-miR-130a-3p to participate in the pathogenesis of CD.AIM To explore the mechanism of hsa_circRNA_102610 in the pathogenesis of CD.METHODS The relative expression levels of hsa_circRNA_102610 and hsa-miR-130a-3p in patients were detected by quantitative reverse transcription-polymerase chain reaction.The proliferation of human intestinal epithelial cells(HIECs)and normal-derived colon mucosa cell line 460(NCM460)cells was detected by cell counting kit-8,5-ethynyl-2’-deoxyuridine staining and cell cycle assays following overexpression or downregulation of hsa_circRNA_102610.Cell proliferation assays were performed as described above in a rescue experiment with hsa-miR-130a-3p mimics.The interaction of hsa_circRNA_102610 and hsa-miR-130a-3p was verified by fluorescence in situ hybridization and dual luciferase reporter assays.The relative expression levels of CyclinD1,mothers against decapentaplegic homolog 4(SMAD4),E-cadherin,N-cadherin and Vimentin were detected by western blotting following hsa_circRNA_102610 overexpression,TGF-β1-induced EMT or hsa-miR-130a-3p mimic transfection(in rescue experiments).RESULTS Upregulation of hsa_circRNA_102610 was determined to be positively correlated with elevated fecal calprotectin levels in CD(r=0.359,P=0.007)by Pearson correlation analysis.Hsa_circRNA_102610 promoted the proliferation of HIECs and NCM460 cells,while hsa-miR-130a-3p reversed the cell proliferationpromoting effects of hsa_circRNA_102610.Fluorescence in situ hybridization and dual luciferase reporter assays showed that hsa_circRNA_102610 directly bound hsa-miR-130a-3p in NCM460 and 293T cells.An inverse correlation between downregulation of hsa-miR-130a-3p and upregulation of hsa_circRNA_102610 in CD patients was observed(r=-0.290,P=0.024)by Pearson correlation analysis.Moreover,overexpression of hsa_circRNA_102610 promoted SMAD4 and CyclinD1 protein expression validated by western-blotting.Furthermore,overexpression of hsa_circRNA_102610 promoted TGF-β1 induced EMT in HIECs and NCM460 cells via targeting of hsa-miR-130a-3p,with increased expression of Vimentin and N-cadherin and decreased expression of E-cadherin.CONCLUSION Hsa_circRNA_102610 upregulation in CD patients could promote the proliferation and EMT of intestinal epithelial cells via sponging of hsa-miR-130a-3p.

Alendronate disturbs femoral growth due to changes during immunolocalization of transforming growth factor-β1 and bone morphogenetic protein-2 in epiphyseal plate

BACKGROUND The epiphyseal growth plate is an important anatomical segment localized on the ends of a long bone.Despite the abovementioned atractive reasons for alendronate’s use,few data on the effect of alendronate during epiphyseal growth exist.AIM Verify the effect of alendronate on the growth epiphyseal plate,and compare its effect with the size of the femur during the double-staining of the immunolocalization of transforming growth factor-β1(TGF-β1)and bone morphogenetic protein-2(BMP2)in endochondral ossifing in specimens that have received alendronate.METHODS Forty newborn rats were randomly divided into two groups:a control group(were given applications of 1 mg/kg physiologic saline)and a group that received Alendronate(a dose of 2.5 mg/kg).These groups were then divided into two subgroups for euthanasia in two and 12 d of life.After euthanasia,the femurs were removed,and the femoral bones were measured linearly between the apex of the greater trochanter until the lower intercondylar midlle face to verify the probable bone growth between 3 and 12 d in control and alednroanto treated rats.Posteriorly,the surgical pieces were also sent to the histopathology laboratory to produce histological slides.The obtained slides were stained with hematoxylin and eosin to measure each of the cartilage zones in endochondral development.and other slides were immunohistochemically tested for anti-TGF-β1 and BMP-2 antibodies to investigate the immunolocalization of these proteins in the epiphyseal plaque area.RESULTS On the third day,some diferences between the control group and specimens treated with alendronate were verified.Macroscopiccaly,we found similarities in size between the femoral bones when we compared the control group with the specimens that received alendronate.On the 12^th day,the bone size of the mice receiving the drug was significantly smaller than those of the control group.These results coincide with changes in the TGF-β1 and BMP-2 expression.In the specimens that received alendronate,the TGF-β1 was expressed in some sites of trabecular bone that was neoformed,peripherally to the bone marrow area.The BMP-2 was also positive in proliferative chondrocytes and hypertrofic chondrocytes.On the 12^th day,all layers of chondrocytes exhibited positivity for BMP-2 in the specimens that received alendronate.In the interface between the trabecular bone and cartilage,an area of disorganized bone deposition was evident.Neoformed bone also appeared to be different at 12 d.In the control group,BMP-2 was positive in an intense area of bone trabeculae,whereas the alendronate-treated group showed TGF-β1 positive trabeculae and a greater bone area.CONCLUSION Alendronate alters the immunolocalization of TGF-β1 and BMP-2 simultaneously,a condition that changes the usual histological aspects of the cartilage zone and impairs epiphysis growth and femur growth.

EMT、TGF-β_1、Ang Ⅱ与器官纤维化发生机制的研究进展

纤维化是大多数慢性炎症性疾病的病理转归,几乎能发生在身体的每个组织器官。纤维化以过多的细胞外基质沉积为特征,进一步发展可导致器官功能衰竭乃至死亡。关于器官纤维化的研究很多,但其确切机制目前尚不明确。近年来,上皮间质转化(EMT)、转化生长因子β1(TGF-β1)、血管紧张素Ⅱ(AngⅡ)在组织器官纤维化形成机制研究中备受关注。该文就EMT、TGF-β1、AngⅡ与各器官纤维化的相互关系及作用机制予以综述,以更全面地认识纤维化的发生机制。

TGF-β_1抗体对体外培养翼状胬肉上皮细胞增殖抑制作用实验研究

目的:探讨不同浓度TGF-β1抗体对翼状胬肉上皮细胞增殖的抑制作用。方法:通过组织块培养法进行人翼状胬肉上皮细胞的体外培养,以不同浓度TGF-β1抗体(0、6.25、12.5、25.0、50.0、100μg/ml)作用于传代培养的翼状胬肉上皮细胞,MTT法检测不同时期(24h、48h、72h)细胞增殖抑制情况;免疫组织化学SABC法染色检测增殖细胞核抗原(PCNA)的表达。结果:组织块培养法成功进行人翼状胬肉上皮细胞的原代和传代培养,并经角蛋白染色鉴定为上皮细胞;不同浓度TGF-β1抗体可明显抑制翼状胬肉上皮细胞的生长,且随浓度增加、时间延长,抑制率明显增高,PCNA的表达不同程度下降,各组间比较有统计学意义。结论:不同浓度TGF-β1抗体可明显抑制翼状胬肉上皮细胞生长,可作为治疗翼状胬肉的新型药物的研究方向。