Dexamethasone alters epithelium proliferation and survival and suppresses Wnt/β-catenin signaling in developing cleft palate

Dexamethasone (Dex) contributes to a cleft palate, but the cellular and molecular mechanisms responsible for the deleterious effect on the developing palate are unclear. Wnt signaling is a causal mechanism of Dex-induced osteoporosis, so this study was conducted to determine whether Dex-induced cleft palate may result from altered Wnt signaling. Administration of Dex to mice completely inhibited canonical Wnt/β-catenin signaling and altered cell proliferation and apoptosis of the craniofacial epithelium in developing embryos. Thus, downregulated Wnt/β-catenin signaling was associated with Dex-induced cleft palate. Moreover, altered cell fate by Dex responsible for small palates, delaying shelf elevation and unfused palates was a crucial mechanism in cleft palate. Our findings help in elucidating the mechanisms of Dex-induced cleft palate.     

Ubiquitination and regulation of Smad7 in the TGF-β1/Smad signaling of aristolochic acid nephropathy

Abstract

Aristolochic acid I (AAI) affects TGF-β1/Smad signaling, which causes AA nephropathy (AAN), but the mechanisms are not fully understood. We aimed to clarify whether Arkadia and UCH37 participate in TGF-β1/Smad signaling via Smad7, and the regulatory mechanisms of Smad7. One side, mice and cultured mouse renal tubular epithelial cells (RTECs) were treated with various AAI doses and concentrations, respectively; on the other side, RTECs were transfected with small interfering RNA (siRNA) expression vectors against Arkadia and UCH37 and then treated with 10?µg/ml AAI. And then detect the mRNA and protein levels of Smad7, UCH37, Arkadia and any other relative factors by RT-PCR and Western blotting. In kidney tissues and RTECs, the mRNA and protein levels of Smad7 decreased with increasing AAI doses concentrations by real-time PCR and Western blotting, whereas those of Arkadia, UCH37, Smad2, Smad3 and TβRI increased. Cells transfected with the Arkadia siRNA expression vector showed reduced mRNA and protein levels of vimentin, α-SMA, Smad2, Smad3 and TβRI after AAI treatment, while those of CK18 and Smad7 increased compared with those of untransfected RTECs. Conversely, cells transfected with the UCH37 siRNA expression vector showed the opposite effect on analyzed signaling molecules after AAI treatment. Arkadia and UCH37 participate in TGF-β1/Smad signaling-mediated renal fibrosis, and Smad7 blocks TGF-β1 signaling by inhibiting Smad2/Smad3 phosphorylation and enhancing the degradation of TβRI.     

Darbepoetin inhibits proliferation of hepatic cancer cells in the presence of TGF-β

Darbepoetin (DPO), an erythropoietin (EPO) derivative, was licensed in 2002 to treat patients with solid tumors suffering from chemotherapy-dependent anemia, although various tumors express EPO to improve vascularization, thus favoring tumor growth and spreading. Therefore, we wanted to investigate direct effects of DPO on the liver tumor cell lines HepG2, SkHep1, Huh-7, AKN1, HCC-T and HCC-M, as well as on primary human hepatocytes (hHeps). DPO (0–40 ng/ml) did not affect viability of hHeps, HepG2, SkHep1, AKN1, HCC-T and HCC-M cells, as determined by Resazurin conversion. However, Huh-7 cells’ viability dose-dependently decreased from 5 ng/ml DPO on. Lack of LDH release into culture medium and negative DNA laddering excluded apoptosis or necrosis as the cause for the reduced Resazurin conversion. In Huh-7 cells, DPO increased the expression of p53. Interestingly, Huh-7 cells showed the highest basal TGF-β₁ expression as compared to the other cell lines. Upon inhibition of TGF-β₁ signaling, DPO no longer reduced viability in Huh-7 cells. On the contrary, co-incubation with TGF-β₁ made the other cell lines responsive to DPO. Summarizing our data, we show that DPO reduces the growth of Huh-7 cells by up-regulation of the tumor-suppressor gene p53. This mechanism seems to be dependent on a strong TGF-β expression and corresponding signaling in these cells, as other cell lines became responsive to DPO with TGF-β₁ supplementation. The knowledge of this mechanism offers great perspectives for the understanding and treatment of solid liver tumors.     

Neferine attenuates development of testosterone-induced benign prostatic hyperplasia in mice by regulating androgen and TGF-β/Smad signaling pathways

Benign prostatic hyperplasia (BPH) is a common urinary disease among the elderly, characterized by abnormal prostatic cell proliferation. Neferine is a dibenzyl isoquinoline alkaloid extracted from Nelumbo nucifera and has antioxidant, anti-inflammatory and anti-prostate cancer effects. The beneficial therapeutic effects and mechanism of action of neferine in BPH remain unclear.A mouse model of BPH was generated by subcutaneous injection of 7.5 mg/kg testosterone propionate (TP) and 2 or 5 mg/kg neferine was given orally for 14 or 28 days. Pathological and morphological characteristics were evaluated. Prostate weight, prostate index (prostate/body weight ratio), expression of type Ⅱ 5α-reductase, androgen receptor (AR) and prostate specific antigen were all decreased in prostate tissue of BPH mice after administration of neferine. Neferine also downregulated the expression of pro-caspase-3, uncleaved PARP, TGF-β1, TGF-β receptor Ⅱ (TGFBR2), p-Smad2/3, N-cadherin and vimentin. Expression of E-cadherin, cleaved PARP and cleaved caspase-3 was increased by neferine treatment.1–100 μM neferine with 1 μM testosterone or 10 nM TGF-β1 were added to the culture medium of the normal human prostate stroma cell line, WPMY-1, for 24 h or 48 h. Neferine inhibited cell growth and production of reactive oxygen species (ROS) in testosterone-treated WPMY-1 cells and regulated the expression of androgen signaling pathway proteins and those related to epithelial-mesenchymal transition (EMT). Moreover, TGF-β1, TGFBR2 and p-Smad2/3, N-cadherin and vimentin expression were increased but E-cadherin was decreased after 24 h TGF-β1 treatment in WPMY-1 cells. Neferine reversed the effects of TGF-β1 treatment in WPMY-1 cells. Neferine appeared to suppress prostate growth by regulating the EMT, AR and TGF-β/Smad signaling pathways in the prostate and is suggested as a potential agent for BPH treatment.     

The role of TGF-β1/Smad3 signaling pathway and oxidative stress in the inhibition of osteoblast mineralization by copper chloride

To explore the relationship of oxidative stress and TGF-β 1/Smad3 pathway in the inhibition of osteoblast mineralization by copper chloride (CuCl₂), the osteoblasts were treated with CuCl₂ (0, 50 μM, 100 μM, 150 μM CuCl₂ 5H₂O) for 24 h. We found that Cu impaired the osteoblast structure, inhibited the glutathione peroxidase (GSH-Px) and superoxide dismutase (SOD) activities, alkaline phosphatase (ALP) content, mRNA expression of collagen I (COL-I), osteocalcin (OCN), insulin-like growth factor I (IGF-I), bone morphogenetic protein-2 (BMP-2), transforming growth factor β1 (TGF-β1) and core-binding factor α1 (Cbfα1), promoted the reactive oxygen species (ROS) production, inactivated the TGF-β1/Smad3 pathway. It indicates that the inactivated TGF-β1/Smad3 pathway leads to osteoblast impairment by CuCl₂. It will contribute to clarify the influence of CuCl₂ on the osteoblast mineralization.     

Strain-dependent effects of transforming growth factor-β1 and 2 during mouse secondary palate development

Cleft palate is a common birth defect affecting 1 in 700 births. Transforming growth factor-βs (TGF-βs) are important signaling molecules, and their functions in murine palate development have received great attention. TGF-β3 is expressed exclusively in palatal epithelial cells and mediates epithelial fusion, whereas the importance of TGF-β1 and 2 in palate have not yet been demonstrated in vivo, since inactivation of Tgf-β1 or Tgf-β2 genes in mice did not reveal significant palate defects. We hypothesized that TGF-β1 and TGF-β2 can compensate each other during palate formation. To test this, we generated Tgf-β1 and Tgf-β2 compound mutant mice and found that approximately 40% of [Tgf-β1^+/−; Tgf-β2^−/−] compound mutant embryos display cleft palate on C57 background. In addition, 26% of Tgf-β2^−/− embryos on 129 background, but not in C57 or Black Swiss, displayed cleft palate. TGF-β1 and 2 functions are required for murine palate development in strain-dependent manner.     

Wen-pi-tang-Hab-Wu-ling-san reduces ureteral obstructive renal fibrosis by the reduction of oxidative stress,inflammation, and TGF-β/Smad2/3 signaling

Kidney fibrosis results in chronic renal disease. The current treatment of chronic renal diseases is limited to angiotensin converting enzyme inhibitors and angiotensin receptor blockers. Recently, we found that Wen-pi-tang-Hab-Wu-ling-san (WHW) extract, which has been used to treat renal diseases in herbal medicine for a long time, plays anti-fibrogenic. Here, we investigated the role of WHW in the kidney fibrosis induced by unilateral ureteral obstruction (UUO) in mice. C57BL/6 male mice were subjected to UUO on day 0 and then administered with either WHW (2, 10, or 50 mg/kg of body weight) or vehicle orally from 1 day after UUO to finish the experiment. WHW-administration significantly mitigated the UUO-induced kidney fibrotic changes including tubular atrophy and dilatation, collagen accumulation, expansion of interstitial space and leukocyte infiltration. WHW prevented the increases of oxidative stress by the prevention of UUO-induced decreases of catalase, copper–zinc superoxide dismutase (CuZnSOD) and manganese superoxide dismutase (MnSOD), resulting in reduced production of oxidative stress. Furthermore, WHW reduced transforming growth factor-β (TGF-β) expression and phosphorylation of Smad2/3 stimulated by UUO. In conclusion, WHW prevented kidney fibrosis following UUO by the inhibition of inflammation, oxidative stress and TGF-β/Smad2/3 signaling pathway.     

Copper oxide nanoparticles induce collagen deposition via TGF-β1/Smad3 signaling in human airway epithelial cells

Use and application of nanoparticles has increased in recent years. Copper oxide nanoparticles (CuONPs) are one of the most common types of nanoparticles, and they are mainly used as catalysts and preservatives. However, limited toxicity data are available on the toxicity of CuONPs to the respiratory system. We investigated fibrotic responses induced by CuONPs in the respiratory tract and elucidated its underlying mechanism of action in vivo and in vitro experiments. In the mouse model, CuONPs exposure markedly increased transforming growth factor-β1 (TGF-β1) and collagen I expression and Smad3 phosphorylation, combined with elevation of inflammatory mediators including interleukin (IL)-1β, IL-6, and tumor necrosis factor-α (TNF-α). These alterations were also observed in histological analysis of lung tissue. CuONPs markedly increased inflammatory responses and collagen deposition, accompanied by the elevation of TGF-β1 and collagen I expression in lung tissue. In addition, CuONPs-treated H292 cells showed significantly increased mRNA and protein production of TGF-β1, collagen I, IL-6, and TNF-α; this response was markedly decreased by treatment of a TGF-β1 inhibitor (SB-431542). Taken together, CuONPs induced fibrotic responses in the respiratory tract, closely related to TGF-β1/Smad3 signaling. Therefore, our results raise the necessity of further investigation for the present state of its risk by providing useful information of the toxicity of CuONPs.     

Macrophages,TGF-β1 expression and iron deposition in development of NASH

A wide range of molecular markers and different types of cells in liver are possible factors for progression of non-alcoholic fatty liver disease (NAFLD), non-alcoholic steatohepatitis (NASH) development of liver fibrosis. We investigated biopsies from 57 patients with NASH. The material was obtained from livers and was proceed immunohistochemistry antibodies against CD68 and TGF-beta 1. In addition, biopsies were evaluated for iron content. Macrophages/-positive/could be found in all 57 cases. The number of macrophages in the sinusoids correlated with the degree of portal fibrosis:64.% of the patients with mild or intensive fibrosis had high infiltration with CD68-positive cells, while 100% of the patients without fibrosis hadlow infiltration (χ²=8.56; p=0.003). In specimens we, 69.% of patients with different degree of fibrosis expressed TGF-β1 in their portal tracts, and 100% of patients without fibrosis did demonstrate expression of the protein (χ²=23.7; p<0.001). Hepatic iron was found in 100% (9) of patients with intensive fibrosis vs. 10.3% of the patients mild fibrosis (χ²=23.4; p<0.001). Our results suggest that the macrophages and macrophage-derived TGF-beta1 are the major factors responsible for development of fibrosis and progression of chronic liver disease.     

Retinoic acid remodels extracellular matrix (ECM) of cultured human fetal palate mesenchymal cells (hFPMCs) through down-regulation of TGF-β/Smad signaling

The regulation of extracellular matrix (ECM) by retinoic acid (RA) is interesting in light of the fact that the ECM plays an essential role in morphogenesis and palatal shelf elevation. In the current study, we explored the effect of RA overexposure on ECM and the probable mechanisms in cultured human fetal palate mesenchymal cells (hFPMCs). RA dose-dependently inhibited cell proliferation and mRNA and protein levels of ECM components fibronectin, tenascin C and fibrillin-2. Zymography revealed that MMP-2 activity was suppressed by RA. Further analysis revealed that mRNA levels of MMP2 and TIMP2 were decreased, while the MMP2/TIMP2 mRNA ratio was increased, which might facilitate the ECM degradation. Because of the pivotal role of TGF-β/Smad pathway in palatogenesis we therefore checked the effect of RA on TGF-β/Smad signaling. The results indicated RA treatment increased Smad7 expression and decreased the levels of TGF-β1, TGF-β3, TGF-β type II receptor (TβRII) and phosphorylated Smad2 and Smad3. Activation of the Smad pathways by either exogenous TGF-β3 or recombinant adenoviruses for Smad3 attenuated RA-induced inhibition of cell proliferation and ECM components and rescued the RA-altered MMP2/TIMP2 mRNA ratio. In conclusion, these findings suggested that RA overexposure inhibited cell proliferation and disrupted the ECM network through down-regulation of TGF-β/Smad pathway.     

Effects of amlodipine on TGF-β-induced Smad2, 4 expressions in adriamycin toxicity of rat mesangial cells

Transforming growth factor-β (TGF-β) is closely associated with progressive renal fibrosis. A central component of TGF-β-stimulated mesangial cell fibrogenesis is the TGF-β family-specific Smad signal transduction pathway. This study investigated the expression of TGF-β-receptor–activated Smad2, its common partner Smad4, and the phosphorylated Smad2 (p-Smad2) in adriamycin-induced toxicity of cultured rat mesangial cells. This in vitro study showed that amlodipine (10⁻⁹ to 10⁻⁵ mol/l) had no effect on the toxicity of rat mesangial cells induced by adriamycin in the absence of TGF-β1. However, amlodipine (10⁻⁷ to 10⁻⁵ mol/l) reduced the toxicity of rat mesangial cells induced by TGF-β1 in the absence of adriamycin; moreover, amlodipine (10⁻⁸ to 10⁻⁵ mol/l) significantly reduced adriamycin-induced cytotoxicity when it was given in combination with TGF-β1; amlodipine (10⁻⁶, 10⁻⁵ mol/l) had no effect on Smad2 mRNA and protein expression induced by adriamycin + TGF-β1, but it (10⁻⁶, 10⁻⁵ mol/l) dramatically inhibited the down-regulation of p-Smad2 protein expression as well as Smad4 mRNA and protein expression induced by adriamycin + TGF-β1 in rat mesangial cells. Present study shows that amlodipine exerts a significant inhibition on adriamycin-induced toxicity in rat mesangial cells by affecting the expression of TGF-β/Smad signaling intermediates p-Smad2 and Smad4.     

Copper nanoparticles induce early fibrotic changes in the liver via TGF-β/Smad signaling and cause immunosuppressive effects in rats

Copper nanoparticles (Cu NPs) have various uses, including as additives in polymers/plastics, lubricants for metallic coating, and biomedical applications. We investigated the role of transforming growth factor (TGF)-β1 signaling in hepatic damage caused by Cu NPs and explored the effects of a 28-day repeated oral administration to Cu NPs on the immune response. The exposure to Cu NPs caused a dose-dependent increase in Cu levels in the liver and spleen. Cu NPs caused hepatic damage and markedly increased oxidative stress in liver tissues. Cu NPs induced activation of TGF-β1/Smad signaling by induction of vascular endothelial growth factor and matrix metalloproteinase-9. Exposure to Cu NPs also induced activation of Smad-independent pathways, phosphorylation of mitogen-activated protein kinases (MAPKs) and Akt/FoxO3. Consistent with the activation of TGF-β1/Smad-dependent and -independent pathways, Cu NPs markedly increased the deposition and induction of extracellular matrix components, α-smooth muscle actin, and collagens in liver tissues. In addition, repeated exposure to Cu NPs suppressed the proliferation of mitogenically stimulated T- or B-lymphocytes and decreased CD3⁺ (particularly, CD3⁺CD4⁺CD8^?) and CD45⁺ population, followed by decreased levels of immunoglobulins and Th1/Th2 type cytokines. Collectively, Cu NPs caused hepatic damage and induced pro-fibrotic changes, which were closely related to the activation of oxidative stress-mediated TGF-β1/Smad-dependent and -independent pathways (MAPKs and Akt/FoxO3). We confirmed the immunosuppressive effect of Cu NPs via the inhibition of mitogen-stimulated spleen-derived lymphocyte proliferation and suppression of B- or T-lymphocyte-mediated immune responses.     

GSK-3β suppresses the proliferation of rat hepatic oval cells through modulating Wnt/β-catenin signaling pathway

Aim:

Glycogen synthase kinase 3β (GSK-3β) plays a crucial role in hepatic biology, including liver development, regeneration, proliferation and carcinogenesis. In this study we investigated the role of GSK-3β in regulation of growth of hepatic oval cells in vitro and in liver regeneration in partially hepatectomized rats.

Methods:

WB-F344 cells, the rat hepatic stem-like epithelial cells, were used as representative of oval cells. Cell viability was examined using a WST-8 assay. The cells were transfected with a recombinant lentivirus expressing siRNA against GSK-3β (GSK-3βRNAiLV) or a lentivirus that overexpressed GSK-3β (GC-GSK-3βLV). Adult rats underwent partial (70%) hepatectomy, and liver weight and femur length were measured at d 7 after the surgery. The expression of GSK-3β, phospho-Ser⁹-GSK-3β, β-catenin and cyclin D1 was examined with immunoblotting assays or immunohistochemistry.

Results:

Treatment of WB-F344 cells with the GSK-3β inhibitor SB216763 (5 and 10 μmol/L) dose-dependently increased the levels of phospho-Ser⁹-GSK-3β, but not the levels of total GSK-3β, and promoted the cell proliferation. Knockout of GSK-3β with GSK-3βRNAiLV increased the cell proliferation, whereas overexpression of GSK-3β with GC-GSK-3βLV decreased the proliferation. Both SB216763 and GSK-3βRNAiLV significantly increased the levels of β-catenin and cyclin D1 in the cells, whereas GSK-3β overexpression decreased their levels. In rats with a partial hepatectomy, administration of SB216763 (2 mg/kg, ip) significantly increased the number of oval cells, the levels of phospho-Ser⁹-GSK-3β, β-catenin and cyclin D1 in liver, as well as the ratio of liver weight to femur length at d 7 after the surgery.

Conclusion:

GSK-3β suppresses the proliferation of hepatic oval cells by modulating the Wnt/β-catenin signaling pathway.     

α-Lipoic acid inhibits liver fibrosis through the attenuation of ROS-triggered signaling in hepatic stellate cells activated by PDGF and TGF-β

Reactive oxygen species (ROS) have been implicated in hepatic stellate cell activation and liver fibrosis. We previously reported that α-lipoic acid (LA) and its reduced form dihydrolipoic acid (DHLA) inhibited toxicant-induced inflammation and ROS generation. In the present study, we further examined the effects of LA/DHLA on thioacetamide (TAA)-induced liver fibrosis in rats and the possible underlying mechanisms in hepatic stellate cells in vitro. We found that co-administration of LA to rats chronically treated with TAA inhibited the development of liver cirrhosis, as indicated by reductions in cirrhosis incidence, hepatic fibrosis, and AST/ALT activities. We also found that DHLA inhibited TGF-β/PDGF-stimulated HSC-T6 activation and ROS generation. These effects could be mediated by the MAPK and PI3K/Akt pathways. According to our current results, LA may have a beneficial role in the treatment of chronic liver diseases caused by ongoing hepatic damage.     

Protocatechuic acid inhibits TGF-β1-induced proliferation and migration of human airway smooth muscle cells

Protocatechuic acid (3, 4-dihydroxybenzoic acid, PCA) is a major metabolite of anthocyanins and was reported to possess anti-allergic response. However, the effects of PCA on airway smooth muscle cells (ASMCs) proliferation and migration remain unclear. Therefore, this study aims to investigate the effects of PCA on proliferation and migration of ASMCs. ASMCs were pre-incubated with various concentrations of PCA for 30 min before stimulation with transforming growth factor-β1 (TGF-β1) for different times. Cell proliferation was determined using the colony formation assay. Cell migration was detected using the Transwell chamber assay. The levels of type I collagen, fibronectin, phosphorylated Smad2, Smad2, phosphorylated Smad3 and Smad3 were detected by western blot analysis. Our results demonstrated that PCA inhibited the proliferation and migration of ASMCs, as well as suppressed the expression levels of type I collagen and fibronectin in ASMCs induced by TGF-β1. Furthermore, PCA obviously down-regulated the phosphorylation levels of Smad2/3 in ASMCs exposed to TGF-β1. Taken together, the present results have revealed that PCA inhibits asthma airway remodeling by suppressing proliferation and extracellular matrix (ECM) protein deposition in TGF-β1-mediated ASMCs via the inactivation of Smad2/3 signaling pathway. Therefore, PCA may be useful for the prevention or treatment of asthma airway remodeling.     

Destruction of the blood-retina barrier in diabetic retinopathy depends on angiotensin-converting enzyme-mediated TGF-β1/Smad signaling pathway activation

Diabetes mellitus (DM) is a systemic, chronic metabolic disease that is related to heredity and autoimmunity and is often accompanied by complications of retinopathy. However, the causative mechanism involved in the pathological process remains unclear. In this research, treatment with fosinopril or LY2109761 was found to be responsible for the improvement of the pathological processes, serum biochemical indexes and retinopathy in rats with streptozotocin-induced diabetes. In addition, the upregulation of angiotensin-converting enzyme (ACE) in the serum and the increased expression of TGF-β1 in the pathological outer nuclear layer (ONL) and inner nuclear layer (INL) of the retina were also reduced. In vitro experiments demonstrated that ACE enhanced cell damage and TGF-β1/Smad signaling pathway activation in retinal capillary endothelial cells (RCECs) under high glucose conditions. In addition, the activity of ACE was also considered to be related to the increasing levels of activated TGF-β1 in both rat retinal Müller cells (RMCs) and RCECs, but ACE activity had no effect on the high glucose-mediated upregulation of total TGF-β1 in RMCs. Coculture experiments with RCECs and RMCs showed that the barrier that was established under normal conditions was significantly impaired when exposed to high glucose combined with ACE, and damage of barrier can be prevented by adding fosinopril or LY2109761. Finally, a similar auxiliary effect of ACE was also observed in the activated TGF-β1-mediated barrier damage in blood-retinal barrier model in vitro. In summary, ACE-mediated TGF-β1/Smad signaling pathway activation was found to be involved in the destruction of the blood-retina barrier during diabetic retinopathy in a model of streptozotocin-induced diabetes, and these data may provide evidence to guide the treatment of the complications of diabetes mellitus.     

Role of TGF-β/Smad signaling pathway in pulmonary fibrosis after rheumatoid arthritis in mice北大核心CSCD

Aim To investigate the role of TGF-β/Smad signaling pathway in rheumatoid arthritis (RA) - associated postinterstitial pulmonary fibrosis in mice. Methods The mouse model of RA was constructed by subcutaneous administration of complete Freund's adjuvant (CFA) and chicken II collagen (Col-II) to the tail root of mice. The blank group was given the same amount of distilled water, and the control group was given the same amount of glacial acetic acid (solvent). The degree of toe swelling (joint swelling degree and arthritis index) was monitored to evaluate the mouse modeling. The pathological changes of mouse lung tissues were observed by HE and Masson staining. The expression of TGF-β in lung tissues were observed by immunohistochemical staining. The level of hydroxyproline in lung tissues was measured by chemiluminescence method. The expressions of Smad2, Smad3 and phosphorylated p-Smad2 and phosphorylated p-Smad3 in lung tissues were detected by Western blot. Results Compared with blank group and solvent group, the joint swelling and arthritis index of model group significantly increased. Twenty-one days after administration, HE staining showed inflammatory changes in lung interstitium of the model group, Masson staining showed collagen fiber deposition and obvious fibrosis in lung interstitium of the model group, and immunohistochemical staining showed that the expression of TGF-β in cytoplasm of lung interstitial cells of the model group increased, which was brown and yellow. Meanwhile, hydroxyproline was significantly raised in lung tissue homogenate of the model group. Further WB analysis showed that compared with blank group and solvent group, the expression of p-Smad2 and pSmad3 in lung tissues of the model group was significantly up-regulated (P < 0. 05, P < 0. 01). Conclusions RA can give rise to pulmonary fibrosis, and the expressions of p-Smad2 and p-Smad3 are up-regulated, which is be pivotal in pulmonary fibrosis and RA-related post-interstitial pulmonary fibrosis. © 2023 Publication Centre of Anhui Medical University. All rights reserved.     

Impairment of ovarian follicular development caused by titanium dioxide nanoparticles exposure involved in the TGF-β/BMP/Smad pathway

Titanium dioxide nanoparticles (TiO₂ NPs) have been shown to induce reproductive system damages in animals. To better underline how TiO₂ NPs act in reproductive system, female mice were exposed to 2.5, 5, or 10 mg/kg TiO₂ NPs by gavage administration for 60 days, the ovary injuries, follicle stimulating hormone (FSH) and luteinizing hormone (LH) levels as well as ovarian follicular development-related molecule expression were investigated. The results showed that TiO₂ NPs exposure resulted in reduction of ovary weight and inhibition of ovarian follicular development. Furthermore, the suppression of follicular development was demonstrated to be closely related to higher FSH and LH levels, and higher expression of activin, follistatin, BMP2, BMP4, TGF-β1, Smad2, Smad3, and Smad4 as well as decreased inhibin-α expression in mouse ovary in a dose-dependent manner. It implies that the impairment of ovarian follicular development caused by TiO₂ NPs exposure may be mediated by TGF-β signal pathway.     

CCM111 prevents hepatic fibrosis via cooperative inhibition of TGF-β, Wnt and STAT3 signaling pathways

CCM111 is an aqueous extract of Antrodia cinnamomea (AC) that has exhibited anti-liver fibrosis functions. However, the detailed mechanisms of AC action against liver fibrosis have not been elucidated yet. The present research showed that CCM111 significantly lowered the levels of the hepatic enzyme markers glutamate oxaloacetate transaminase (GOT) and glutamic pyruvic transaminase (GPT), prevented liver damage and collagen deposition, and downregulated TGF-β/Smad signaling in a dose-dependent manner compared with CCl₄ treatment alone. CCM111 markedly inhibited TGF-β, Wnt and STAT3 signaling pathway-regulated downstream genes in the liver by next-generation sequencing. The antifibrotic mechanisms of CCM111 were further demonstrated in HSC-T6 cells. Our data demonstrated for the first time that CCM111 can protect against CCl₄-induced liver fibrosis by the cooperative inhibition of TGF-β-, Wnt- and STAT3-dependent proinflammatory and profibrotic mediators, suggesting that CCM111 might be a candidate for preventing and treating chronic fibrotic liver diseases.