KAI1/CD82 decreases Rac1 expression and cell proliferation through PI3K/Akt/mTOR pathway in H1299 lung carcinoma cells

Although the KAI1/CD82 protein has been reported to inhibit cell metastasis in many studies, its mechanism of action has not yet been fully elucidated. In the present study, we investigated the possible effects of KAI1/CD82 on the metastatic phenotype in H1299 lung carcinoma cells. These studies were based on the pivotal role that the acquisition of motile phenotype plays on the initial steps of metastasis. KAI1/CD82‐mediated morphological changes were observed using phase contrast microscopy. We report here, that a KAI1/CD82‐induced phenotypic change was involved in the decrease of Rac1 expression and GTPase activity. However, we found that KAI1/CD82 did not regulate Rac1 mRNA levels. This suggests the existence of another regulatory mechanism of Rac1 protein maturation or activation. To identify the signaling pathway of Rac1 regulation, we investigated the PI3K/Akt/mTOR pathway, since the PI3K/Akt pathway regulates Rac1 activation and mTOR is known to play a regulatory role in protein translation. H1299/CD82‐transfectants showed lower mTOR expression and cell growth than the control group. The data obtained from this study suggested that KAI1/CD82 decreased the metastatic phenotype of H1299 lung carcinoma cells by down‐regulating Rac1 expression through the PI3K/Akt/mTOR pathway. Copyright © 2008 John Wiley & Sons, Ltd.     

MFTZ‐1 reduces constitutive and inducible HIF‐1α accumulation and VEGF secretion independent of its topoisomerase II inhibition

The macrolide compound MFTZ‐1 has been identified as a novel topoisomerase II (Top2) inhibitor with potent in vitro and in vivo anti‐tumour activities. In this study, we further examined the effects of MFTZ‐1 on hypoxia‐inducible factor‐1α (HIF‐1α) accumulation, vascular endothelial growth factor (VEGF) secretion and angiogenesis. MFTZ‐1 reduced HIF‐1α accumulation driven by hypoxia or growth factors in human cancer cells. Mechanistic studies revealed that MFTZ‐1 did not affect the degradation of HIF‐1α protein or the level of HIF‐1α mRNA. By contrast, MFTZ‐1 apparently inhibited constitutive and inducible activation of both phosphatidylinositol‐3‐kinase (PI3K)‐Akt and p42/p44 mitogen‐activated protein kinase (MAPK) pathways. Further studies revealed that MFTZ‐1 abrogated the HIF‐1α‐driven increase in VEGF mRNA and protein secretion. MFTZ‐1 also lowered the basal level of VEGF secretion. The results reveal an important feature that MFTZ‐1 can reduce constitutive, HIF‐1α‐independent VEGF secretion and concurrently antagonize inducible, HIF‐1α‐dependent VEGF secretion. Moreover, MFTZ‐1 disrupted tube formation of human umbilical vein endothelial cells (HUVECs) stimulated by hypoxia with low‐concentration serum or by serum at normoxia, and inhibited HUVECs migration at normoxia. MFTZ‐1 also prevented microvessel outgrowth from rat aortic ring. These data reflect the potent anti‐angiogenesis of MFTZ‐1 under different conditions. Furthermore, using specific small interfering RNA targeting Top2α or Top2‐defective HL60/MX2 cells, we showed that MFTZ‐1 affected HIF‐1α accumulation and HUVECs tube formation irrelevant to its Top2 inhibition. Taken together, our data collectively reveal that MFTZ‐1 reduces constitutive and inducible HIF‐1α accumulation and VEGF secretion possibly via PI3K‐Akt and MAPK pathways, eliciting anti‐angiogenesis independently of its Top2 inhibition.     

An allelic series at the KARRIKIN INSENSITIVE 2 locus of Arabidopsis thaliana decouples ligand hydrolysis and receptor degradation from downstream signalling

Karrikins are butenolide compounds present in post‐fire environments that can stimulate seed germination in many species, including Arabidopsis thaliana. Plants also produce endogenous butenolide compounds that serve as hormones, namely strigolactones (SLs). The receptor for karrikins (KARRIKIN INSENSITIVE 2; KAI2) and the receptor for SLs (DWARF14; D14) are homologous proteins that share many similarities. The mode of action of D14 as a dual enzyme receptor protein is well established, but the nature of KAI2‐dependent signalling and its function as a receptor are not fully understood. To expand our knowledge of how KAI2 operates, we screened ethyl methanesulphonate (EMS)‐mutagenized populations of A. thaliana for mutants with kai2‐like phenotypes and isolated 13 new kai2 alleles. Among these alleles, kai2‐10 encoded a D184N protein variant that was stable in planta. Differential scanning fluorimetry assays indicated that the KAI2 D184N protein could interact normally with bioactive ligands. We developed a KAI2‐active version of the fluorescent strigolactone analogue Yoshimulactone Green to show that KAI2 D184N exhibits normal rates of ligand hydrolysis. KAI2 D184N degraded in response to treatment with exogenous ligands, suggesting that receptor degradation is a consequence of ligand binding and hydrolysis, but is insufficient for signalling activity. Remarkably, KAI2 D184N degradation was hypersensitive to karrikins, but showed a normal response to strigolactone analogues, implying that these butenolides may interact differently with KAI2. These results demonstrate that the enzymatic and signalling functions of KAI2 can be decoupled, and provide important insights into the mechanistic events that underpin butenolide signalling in plants.     

A protective role of ciglitazone in ox‐LDL‐induced rat microvascular endothelial cells via modulating PPARγ‐dependent AMPK/eNOS pathway

Thiazolidinediones, the antidiabetic agents such as ciglitazone, has been proved to be effective in limiting atherosclerotic events. However, the underlying mechanism remains elucidative. Ox‐LDL receptor‐1 (LOX‐1) plays a central role in ox‐LDL‐mediated atherosclerosis via endothelial nitric oxide synthase (eNOS) uncoupling and nitric oxide reduction. Therefore, we tested the hypothesis that ciglitazone, the PPARγ agonist, protected endothelial cells against ox‐LDL through regulating eNOS activity and LOX‐1 signalling. In the present study, rat microvascular endothelial cells (RMVECs) were stimulated by ox‐LDL. The impact of ciglitazone on cell apoptosis and angiogenesis, eNOS expression and phosphorylation, nitric oxide synthesis and related AMPK, Akt and VEGF signalling pathway were observed. Our data showed that both eNOS and Akt phosphorylation, VEGF expression and nitric oxide production were significantly decreased, RMVECs ageing and apoptosis increased after ox‐LDL induction for 24 hrs, all of which were effectively reversed by ciglitazone pre‐treatment. Meanwhile, phosphorylation of AMP‐activated protein kinase (AMPK) was suppressed by ox‐LDL, which was also prevented by ciglitazone. Of interest, AMPK inhibition abolished ciglitazone‐mediated eNOS function, nitric oxide synthesis and angiogenesis, and increased RMVECs ageing and apoptosis. Further experiments showed that inhibition of PPARγ significantly suppressed AMPK phosphorylation, eNOS expression and nitric oxide production. Ciglitazone‐mediated angiogenesis and reduced cell ageing and apoptosis were reversed. Furthermore, LOX‐1 protein expression in RMVECs was suppressed by ciglitazone, but re‐enhanced by blocking PPARγ or AMPK. Ox‐LDL‐induced suppression of eNOS and nitric oxide synthesis were largely prevented by silencing LOX‐1. Collectively, these data demonstrate that ciglitazone‐mediated PPARγ activation suppresses LOX‐1 and moderates AMPK/eNOS pathway, which contributes to endothelial cell survival and function preservation.     

Glycation of fibronectin inhibits VEGF‐induced angiogenesis by uncoupling VEGF receptor‐2‐c‐Src crosstalk

Glycation of extracellular matrix proteins has been demonstrated to contribute to the pathogenesis of vascular complications. However, no previous report has shown the role of glycated fibronectin (FN) in vascular endothelial growth factor (VEGF)‐induced angiogenesis. Thus, this study aimed to investigate the effects of glycated FN on VEGF signalling and to clarify the molecular mechanisms involved. FN was incubated with methylglyoxal (MGO) in vitro to synthesize glycated FN, and human umbilical vein endothelial cells (HUVECs) were seeded onto unmodified and MGO‐glycated FN. Then, VEGF‐induced angiogenesis and VEGF‐induced VEGF receptor‐2 (VEGFR‐2) signalling activation were measured. The results demonstrated that normal FN‐positive bands (260 kD) vanished and advanced glycation end products (AGEs) appeared in MGO‐glycated FN and glycated FN clearly changed to a higher molecular mass. The glycation of FN inhibited VEGF‐induced VEGF receptor‐2 (VEGFR‐2), Akt and ERK1/2 activation and VEGF‐induced cell migration, proliferation and tube formation. The glycation of FN also inhibited the recruitment of c‐Src to VEGFR‐2 by sequestering c‐Src through receptor for AGEs (RAGE) and the anti‐RAGE antibody restored VEGF‐induced VEGFR‐2, Akt and ERK1/2 phosphorylation, endothelial cell migration, proliferation and tube formation. Furthermore, the glycation of FN significantly inhibited VEGF‐induced neovascularization in the Matrigel plugs implanted into subcutaneous tissue of mice. Taken together, these data suggest that the glycation of FN may inhibit VEGF signalling and VEGF‐induced angiogenesis by uncoupling VEGFR‐2‐c‐Src interaction. This may provide a novel mechanism for the impaired angiogenesis in diabetic ischaemic diseases.     

Tumour cell‐intrinsic CTLA4 regulates PD‐L1 expression in non‐small cell lung cancer

Cytotoxic T lymphocyte antigen 4 (CTLA4) and programmed cell death protein 1 (PD‐1) are immune checkpoint proteins expressed in T cells. Although CTLA4 expression was found in multiple tumours including non‐small cell lung cancer (NSCLC) tissues and cells, its function in tumour cells is unknown. Recently, PD‐1 was found to be expressed in melanoma cells and to promote tumorigenesis. We found that CTLA4 was expressed in a subset of NSCLC cell lines and in a subgroup of cancer cells within the lung cancer tissues. We further found that in NSCLC cells, anti‐CTLA4 antibody can induce PD‐L1 expression, which is mediated by CTLA4 and the EGFR pathway involving phosphorylation of MEK and ERK. In CTLA4 knockout cells, EGFR knockout cells or in the presence of an EGFR tyrosine kinase inhibitor, anti‐CTLA4 antibody was not able to induce PD‐L1 expression in NSCLC cells. Moreover, anti‐CTLA4 antibody promoted NSCLC cell proliferation in vitro and tumour growth in vivo in the absence of adaptive immunity. These results suggest that tumour cell‐intrinsic CTLA4 can regulate PD‐L1 expression and cell proliferation, and that anti‐CTLA4 antibody, by binding to the tumour cell‐intrinsic CTLA4, may result in the activation of the EGFR pathway in cancer cells.     

Effect of the in vivo application of granulocyte colony‐stimulating factor on NK cells in bone marrow and peripheral blood

Granulocyte colony‐stimulating factor (G‐CSF) has been widely used in the field of allogeneic haematopoietic stem cell transplantation (allo‐HSCT) for priming donor stem cells from the bone marrow (BM) to peripheral blood (PB) to collect stem cells more conveniently. Donor‐derived natural killer (NK) cells have important antitumour functions and immune regulatory roles post‐allo‐HSCT. The aim of this study was to evaluate the effect of G‐CSF on donors' NK cells in BM and PB. The percentage of NK cells among nuclear cells and lymphocyte was significantly decreased and led to increased ratio of T and NK cells in BM and PB post‐G‐CSF in vivo application. Relative expansion of CD56^bri NK cells led to a decreased ratio of CD56^dim and CD56^bri NK subsets in BM and PB post‐G‐CSF in vivo application. The expression of CD62L, CD54, CD94, NKP30 and CXCR4 on NK cells was significantly increased in PB after G‐CSF treatment. G‐CSF treatment decreased the IFN‐γ‐secreting NK population (NK1) dramatically in BM and PB, but increased the IL‐13‐secreting NK (NK2), TGF‐β‐secreting NK (NK3) and IL‐10‐secreting NK (NKr) populations significantly in BM. Clinical data demonstrated that higher doses of NK1 infused into the allograft correlated with an increased incidence of chronic graft‐vs‐host disease post‐transplantation. Taken together, our results show that the in vivo application of G‐CSF can modulate NK subpopulations, leading to an increased ratio of T and NK cells and decreased ratio of CD56^dim and CD56^bri NK cells as well as decreased NK1 populations in both PB and BM.     

ZEB1‐mediated vasculogenic mimicry formation associates with epithelial–mesenchymal transition and cancer stem cell phenotypes in prostate cancer

The zinc finger E‐box‐binding homeobox 1 (ZEB1) induced the epithelial–mesenchymal transition (EMT) and altered ZEB1 expression could lead to aggressive and cancer stem cell (CSC) phenotypes in various cancers. Tissue specimens from 96 prostate cancer patients were collected for immunohistochemistry and CD34/periodic acid–Schiff double staining. Prostate cancer cells were subjected to ZEB1 knockdown or overexpression and assessment of the effects on vasculogenic mimicry formation in vitro and in vivo. The underlying molecular events of ZEB1‐induced vasculogenic mimicry formation in prostate cancer were then explored. The data showed that the presence of VM and high ZEB1 expression was associated with higher Gleason score, TNM stage, and lymph node and distant metastases as well as with the expression of vimentin and CD133 in prostate cancer tissues. Furthermore, ZEB1 was required for VM formation and altered expression of EMT‐related and CSC‐associated proteins in prostate cancer cells in vitro and in vivo. ZEB1 also facilitated tumour cell migration, invasion and clonogenicity. In addition, the effects of ZEB1 in prostate cancer cells were mediated by Src signalling; that is PP2, a specific inhibitor of the Src signalling, dose dependently reduced the p‐Src⁵²⁷ level but not p‐Src⁴¹⁶ level, while ZEB1 knockdown also down‐regulated the level of p‐Src⁵²⁷ in PC3 and DU‐145 cells. PP2 treatment also significantly reduced the expression of VE‐cadherin, vimentin and CD133 in these prostate cancer cells. Src signalling mediated the effects of ZEB1 on VM formation and gene expression.     

miR‐148a inhibits early relapsed colorectal cancers and the secretion of VEGF by indirectly targeting HIF‐1α under non‐hypoxia/hypoxia conditions

Vascular endothelial growth factor (VEGF) is correlated with angiogenesis and early relapse of colorectal cancer (CRC). This study investigated the role of miR‐148a in the regulation of VEGF/angiogenesis and early relapse of CRC. We established a stable clone with miR‐148a expression in HCT116 and HT29 cell lines and created a hypoxic condition by using CoCl₂ to determine the underlying mechanism of miR‐148a. The effects of miR‐148a on the phosphoryl‐ERK (pERK)/hypoxia‐inducible factor‐1α (HIF‐1α)/VEGF pathway were evaluated through Western blotting and the inhibitory effect of miR‐148a on angiogenesis was demonstrated through a tube formation assay. Sixty‐three CRC tissues (28 early relapse and 35 non‐early relapse) were analysed to assess the relationship between miR‐148a and HIF‐1α/VEGF. The protein expression of pERK/HIF‐1α/VEGF in HCT116 and HT29 cells was significantly decreased by miR‐148a (all P < 0.05). The protein expression of VEGF/HIF‐1α was strongly inversely associated with the expression of miR‐148a in the 63 CRC tissue samples (all P < 0.05). Tube formation assay demonstrated that miR‐148a significantly obliterated angiogenesis. miR‐148a suppresses VEGF through down‐regulation of the pERK/HIF‐1α/VEGF pathway and might lead to the inhibition of angiogenesis; miR‐148a down‐regulation increased the early relapse rate of CRC. This demonstrates that miR‐148a is a potential diagnostic and therapeutic target.     

CXCR4/CXCL12 axis promotes VEGF-mediated tumor angiogenesis through Akt signaling pathway

CXC chemokine receptor 4 (CXCR4) has been shown to play a critical role in chemotaxis and homing, which are key steps in cancer metastasis. There is also increasing evidence that links this receptor to angiogenesis; however, its molecular basis remains elusive. Vascular endothelial growth factor (VEGF), one of the major angiogenic factors, promotes the formation of leaky tumor vasculatures that are the hallmarks of tumor progression. Here, we investigated whether CXCR4 induces the expression of VEGF through the PI3K/Akt pathway. Our results showed that CXCR4/CXCL12 induced Akt phosphorylation, which resulted in upregulation of VEGF at both the mRNA and protein levels. Conversely, blocking the activation of Akt signaling led to a decrease in VEGF protein levels; blocking CXCR4/CXCL12 interaction with a CXCR4 antagonist suppressed tumor angiogenesis and growth in vivo. Furthermore, VEGF mRNA levels correlated well with CXCR4 mRNA levels in patient tumor samples. In summary, our study demonstrates that the CXCR4/CXCL12 signaling axis can induce angiogenesis and progression of tumors by increasing expression of VEGF through the activation of PI3K/Akt pathway. Our findings suggest that targeting CXCR4 could provide a potential new anti-angiogenic therapy to suppress the formation of both primary and metastatic tumors.     

Feeding transgenic plants that express a tolerogenic fusion protein effectively protects against arthritis

Although much explored, oral tolerance for treatment of autoimmune diseases still awaits the establishment of novel and effective vectors. We investigated whether the tolerogenic CTA1(R7K)‐COL‐DD fusion protein can be expressed in edible plants, to induce oral tolerance and protect against arthritis. The fusion protein was recombinantly expressed in Arabidopsis thaliana plants, which were fed to H‐2^q‐restricted DBA/1 mice to assess the preventive effect on collagen‐induced arthritis (CIA). The treatment resulted in fewer mice exhibiting disease and arthritis scores were significantly reduced. Immune suppression was evident in treated mice, and serum biomarkers for inflammation as well as anticollagen IgG responses were reduced. In spleen and draining lymph nodes, CD4⁺ T‐cell responses were reduced. Concomitant with a reduced effector T‐cell activity with lower IFNγ, IL‐13 and IL‐17A production, we observed an increase in IL‐10 production to recall antigen stimulation in vitro, suggesting reduced Th1, Th2 and Th17 activity subsequent to up‐regulated IL‐10 and regulatory T‐cell (Treg) functions. This study shows that edible plants expressing a tolerogen were effective at stimulating CD4 T‐cell tolerance and in protecting against CIA disease. Our study conveys optimism as to the potential of using edible plants for oral treatment of rheumatoid arthritis.     

Cimifugin suppresses allergic inflammation by reducing epithelial derived initiative key factors via regulating tight junctions

Cimifugin is a bioactive component of Saposhnikovia divaricata, a Chinese herb for treating allergy. Our previous studies demonstrated that cimifugin inhibited allergic inflammation efficiently. This study aims to determine the mechanism of cimifugin on epithelial cells in allergic inflammation. Mice were sensitized and challenged with FITC to establish type 2 atopic dermatitis (AD) model. The initial stage of AD model, in which mice were just sensitized with FITC, was established in vivo and immortalized human epidermal (HaCaT) cells were utilized in vitro. Initiative key cytokines, TSLP and IL‐33, were measured by ELISA, the junctions in ECs were observed by electron microscopy and TJs (CLDN‐1, occludin and CLDND1) were assessed by Western blot, immunohistochemistry and immunofluorescence. The results showed that TSLP and IL‐33 were inhibited significantly by cimifugin in the initial stage of AD model. Simultaneously, cimifugin reduced the separated gap among the epithelial cells and increased the expression of TJs. Similar effects on TSLP/IL‐33 and TJs were obtained in vitro. The effect of cimifugin on TSLP decreased significantly when expression of CLDN1 was interfered with siRNA and this implied cimifugin inhibits initiative cytokines through restoring TJs. Furthermore, cimifugin administered only in the initial stage obviously attenuated the ultimate allergic inflammation, which indicate that impacts of cimifugin in the initial stage on TSLP/IL‐33 and TJs are sufficient for suppressing allergic inflammation. This study not only revealed the mechanisms of cimifugin, but also indicated the possibility of initiative key cytokines and TJs as therapeutic targets.     

CD151 mediates netrin‐1‐induced angiogenesis through the Src‐FAK‐Paxillin pathway

Crosstalk between the nervous and vascular systems is important during development and in response to injury, and the laminin‐like axonal guidance protein netrin‐1 has been studied for its involvement in angiogenesis and vascular remodelling. In this study, we examined the role of netrin‐1 in angiogenesis and explored the underlying mechanisms. The effect of netrin‐1 on brain tissues and endothelial cells was examined by immunohistochemistry and western blotting in a middle cerebral artery occlusion model and in human umbilical vein endothelial cells. Cell proliferation and cell cycle progression were assessed by the MTT assay and flow cytometry, and the Transwell and tube formation assays were used to examine endothelial cell motility and function. Netrin‐1 up‐regulated CD151 and VEGF concomitant with the activation of focal adhesion kinase (FAK), Src and Paxillin in vitro and in vivo and the induction of cell proliferation, migration and tube formation in vitro. Silencing of CD151 abolished the effects of netrin‐1 on promoting cell migration and tube formation mediated by the activation of FAK/Src signalling. Netrin‐1 promoted angiogenesis in vitro and in vivo by activating the FAK/Src/Paxillin signalling pathway through a mechanism dependent on the expression of the CD151 tetraspanin, suggesting the existence of a netrin‐1/FAK/Src/CD151 signalling axis involved in the modulation of angiogenesis.     

Nicotinamide phosphoribosyltransferase (NAMPT/PBEF/visfatin) is a tumoural cytokine released from melanoma

High plasma levels of nicotinamide phosphoribosyltransferase (NAMPT), traditionally considered an intracellular enzyme with a key role in NAD synthesis, have been reported in several oncological, inflammatory and metabolic diseases. We now show that eNAMPT can be actively released by melanoma cells in vitro. We analysed the mechanisms of its release, and we found both classical and non‐classical pathway involvement. eNAMPT released by melanoma cells, in our hands, has paracrine and autocrine effects: it activates MAPK, AKT and NF‐κB pathways and increases colony formation in anchorage‐independent conditions. eNAMPT also induces M1 polarization in human monocytes. Last, we demonstrate, for the first time in any cancer type, that eNAMPT levels in plasma of tumour‐bearing mice increase and that this increase can be reconducted to the tumour itself. This provides an important cue on previous observations that eNAMPT is increased in patients with cancer. Moreover, silencing NAMPT in melanoma cells leads to a reduction in the tumour growth rate. Our findings extend the basis to consider eNAMPT as a cytokine involved in tumour progression.     

Blood vessel formation in cerebral organoids formed from human embryonic stem cells

Current cerebral organoid technology provides excellent in vitro models mimicking the structure and function of the developing human brain, which enables studies on normal and pathological brain; however, further improvements are necessary to overcome the problems of immaturity and dearth of non-parenchymal cells. Vascularization is one of the major challenges for recapitulating processes in the developing human brain. Here, we examined the formation of blood vessel-like structures in cerebral organoids induced by vascular endothelial growth factor (VEGF) in vitro. The results indicated that VEGF enhanced differentiation of vascular endothelial cells (ECs) without reducing neuronal markers in the embryonic bodies (EBs), which then successfully developed into cerebral organoids with open-circle vascular structures expressing an EC marker, CD31, and a tight junction marker, claudin-5, characteristic of the blood-brain barrier (BBB). Further treatment with VEGF and Wnt7a promoted the formation of the outer lining consisting of pericyte-like cells, which surrounded the vascular tubes. RNA sequencing revealed that VEGF upregulated genes associated with tube formation, vasculogenesis, and the BBB; it also changed the expression of genes involved in brain embryogenesis, suggesting a role of VEGF in neuronal development. These results indicate that VEGF treatment can be used to generate vessel-like structures with mature BBB characteristics in cerebral organoids in vitro.     

Interleukin-11 increases cell motility and up-regulates intercellular adhesion molecule-1 expression in human chondrosarcoma cells

Interleukin‐11 (IL‐11) was originally identified as the cytokine that could induce the proliferation of human cells. Recent studies have shown that IL‐11 plays a critical role in tumor growth, angiogenesis, and metastasis. Chondrosarcoma is a type of highly malignant tumor with a potent capacity to invade locally and cause distant metastasis. However, the effects of IL‐11 on human chondrosarcoma cells are largely unknown. Here, we found that IL‐11 increased the migration and expression of intercellular adhesion molecule‐1 (ICAM)‐1 in human chondrosarcoma cells. We also found that human chondrosarcoma tissues had significant expression of the IL‐11 which was higher than that in primary chondrocytes. The phosphatidylinositol 3‐kinase (PI3K), Akt, and NF‐κB pathways were activated by IL‐11 treatment, and the IL‐11‐induced expression of ICAM‐1 and migration activity were inhibited by the specific inhibitors and mutant forms of PI3K, Akt, and NF‐κB cascades. Taken together, our results indicate that IL‐11 enhanced the migration of the chondrosarcoma cells by increasing ICAM‐1 expression through the IL‐11Rα receptor, PI3K, Akt, and NF‐κB signal transduction pathway. J. Cell. Biochem. 113: 3353–3362, 2012. © 2012 Wiley Periodicals, Inc.     

Thyroid hormone induces sprouting angiogenesis in adult heart of hypothyroid mice through the PDGF‐Akt pathway

Study of physiological angiogenesis and associated signalling mechanisms in adult heart has been limited by the lack of a robust animal model. We investigated thyroid hormone‐induced sprouting angiogenesis and the underlying mechanism. Hypothyroidism was induced in C57BL/6J mice by feeding with propylthiouracil (PTU). One year of PTU treatment induced heart failure. Both 12 weeks‐ (young) and 1 year‐PTU (middle age) treatment caused a remarkable capillary rarefaction observed in capillary density. Three‐day Triiodothyronine (T3) treatment significantly induced cardiac capillary growth in hypothyroid mice. In cultured left ventricle (LV) tissues from PTU‐treated mice, T3 also induced robust sprouting angiogenesis where pericyte‐wrapped endothelial cells formed tubes. The in vitro T3 angiogenic response was similar in mice pre‐treated with PTU for periods ranging from 1.5 to 12 months. Besides bFGF and VEGF₁₆₄, PDGF‐BB was the most robust angiogenic growth factor, which stimulated notable sprouting angiogenesis in cultured hypothyroid LV tissues with increasing potency, but had little effect on tissues from euthyroid mice. T3 treatment significantly increased PDGF receptor beta (PDGFR‐β) protein levels in hypothyroid heart. PDGFR inhibitors blocked the action of T3 both on sprouting angiogenesis in cultured LV tissue and on capillary growth in vivo. In addition, activation of Akt signalling mediated in T3‐induced angiogenesis was blocked by PDGFR inhibitor and neutralizing antibody. Our results suggest that hypothyroidism leads to cardiac microvascular impairment and rarefaction with increased sensitivity to angiogenic growth factors. T3‐induced cardiac sprouting angiogenesis in adult hypothyroid mice was associated with PDGF‐BB, PDGFR‐β and downstream activation of Akt.