Leukemia inhibitory factor (LIF) is a member of the IL-6 cytokine family that functions in the survival, repair and formation of neurons as well as in the maintenance of neural and embryonic stem cells. The functions of LIF have been well documented in mammals, however until recently, the presence of IL-6 family cytokines in ectothermic vertebrates has only been speculated. We report on the identification of lif and lifr transcripts in the zebrafish and document the expression of these molecules in the developing embryos and tissues of adult zebrafish. We also examined the phylogenetic relationship between these molecules and other IL-6 cytokine family members known in mammals. In adult zebrafish, lif is expressed in the kidney and brain while lifr is expressed in the kidney, gill, brain, spleen and liver. During zebrafish embryogenesis, lif and lifr are both expressed as early as 12 hours postfertilization (hpf). In developing zebrafish, lif is expressed in the otic vesicle, retina and cranial sensory ganglia, and lifr is strongly expressed in the notochord, forebrain, otic vesicle, cranial ganglia and the retina. Morpholino knockdown of Lif and Lifr in developing embryos suggests that Lifr, but not Lif is required for proper neural development. lifr morpholino-injected embryos exhibit defects in the trigeminal, facial and vagal branchiomotor neurons, and improper axonal development as measured by acetylated tubulin staining. These embryos also display severe hydrocephaly by 48 hpf. This suggests that Lifrs are involved in proper neural development in zebrafish. This is the first evidence of the expression and role of an LIFR-like molecule in developing fish.

The direction of cytokine secretion from polarized cells determines the cytokine's cellular targets. Leukemia inhibitory factor (LIF) belongs to the interleukin-6 (IL-6) family of cytokines and signals through LIFR/gp130. Three factors which may regulate the direction of LIF secretion were studied: the site of stimulation, signal peptides, and expression levels. Stimulation with IL-1beta is known to promote IL-6 secretion from the stimulated membrane (apical or basolateral) in the human intestinal epithelial cell line Caco-2. Since LIF is related to IL-6, LIF secretion was also tested in Caco-2 following IL-1beta stimulation. Signal peptides may influence the trafficking of LIF. Two isoforms of murine LIF, LIF-M and LIF-D, encode different signal peptides which have been associated with different locations of the mature protein in fibroblasts. To determine the effect of the signal peptides on LIF secretion, secretion levels were compared in Madin-Darby canine kidney (MDCK) clones which expressed murine LIF-M or LIF-D or human LIF under the control of an inducible promoter. Low and high levels of LIF expression were also compared since saturation of the apical or basolateral route would reveal specific transporters for LIF.

Despite advances in assisted reproductive technology, treatment for deficient endometrial receptivity is a major clinical unmet need. In our previous study, the water extract of Paeonia lactiflora Pall. enhanced endometrial receptivity in vitro and in vivo via induction of leukemia inhibitory factor (LIF), an interleukin (IL)-6 family cytokine. In the present study, we found that paeoniflorin, a monoterpene glycoside, is the major active compound of P. lactiflora. Paeoniflorin significantly improved the embryo implantation rate in a murine model of mifepristone (RU486)-induced implantation failure. In addition, paeoniflorin increased the adhesion of human trophectoderm-derived JAr cells to endometrial Ishikawa cells through the expression of LIF in vitro. Moreover, using the National Center for Biotechnology Information (NCBI) Gene Expression Omnibus (GEO) database of the human endometrium, we confirmed that LIF signaling is a key regulator for improving human endometrial receptivity. Therefore, these results suggest that paeoniflorin might be a potent drug candidate for the treatment of endometrial implantation failure by enhancing endometrial receptivity.

Leukemia Inhibitory Factor (LIF) is a polyfunctional cytokine, involved in numerous regulatory effects in vivo and in vitro, varying from cell proliferation to differentiation, and has therapeutic potential for treating various diseases. In the current study, a COS-1 cell line overexpressing recombinant Buffalo LIF (rBuLIF) was established. The rBuLIF was purified to homogeneity from the total cell lysate of COS-1 cells using a two-step affinity chromatography. The purified LIF was confirmed by western blot and mass spectrometer (MS/MS). Particularly, high-resolution MS has identified the rBuLIF with 73% of sequence coverage with highest confidence parameters and with the search engine score of 4580. We determined the molecular weight of rBuLIF protein to be 58.99 kDa and 48.9 kDa with and without glycosylation, respectively. Moreover, the purified rBuLIF was verified to be functionally active by measuring the growth inhibition of M1 myeloid leukemia cells, revealing a maximum inhibition at 72 hours and half-maximal effective concentration (EC50) of 0.0555 ng/ml, corresponding to a specific activity of >1.6×10(7) units/mg. Next, we evaluated the effect of rBuLIF on buffalo mammary epithelial cell lines for its role in involution and also identified the IC50 value for BuMEC migrating cells to be 77.8 ng/ml. Additionally, the treatment of MECs (BuMEC and EpH4) displayed significant (P < 0.05) reduction in growth progression, as confirmed by qRT-PCR analysis, suggesting its strong involvement in the involution of the mammary gland in vivo. Thus, we conclude that the glycosylated rBuLIF, purified from COS-1 cells was found to be functionally active as its natural counterpart.

Ectopic pregnancy is unique to humans and a leading cause of maternal morbidity and mortality. The etiology remains unknown however factors regulating embryo implantation likely contribute. Leukemia inhibitory factor (LIF) has roles in extravillous trophoblast adhesion and invasion and is present in ectopic implantation sites. We hypothesised that LIF facilitates blastocyst adhesion/invasion in the Fallopian tube, contributing to ectopic pregnancy.

Cardiac stem cells or precursor cells regenerate cardiomyocytes; however, the mechanism underlying this effect remains unclear. We generated CreLacZ mice in which more than 99.9% of the cardiomyocytes in the left ventricular field were positive for 5-bromo-4-chloro-3-indolyl-β-d-galactoside (X-gal) staining immediately after tamoxifen injection. Three months after myocardial infarction (MI), the MI mice had more X-gal-negative (newly generated) cells than the control mice (3.04 ± 0.38/mm2, MI; 0.47 ± 0.16/mm2, sham; p < 0.05). The cardiac side population (CSP) cell fraction contained label-retaining cells, which differentiated into X-gal-negative cardiomyocytes after MI. We injected a leukemia inhibitory factor (LIF)-expression construct at the time of MI and identified a significant functional improvement in the LIF-treated group. At 1 month after MI, in the MI border and scar area, the LIF-injected mice had 31.41 ± 5.83 X-gal-negative cardiomyocytes/mm2, whereas the control mice had 12.34 ± 2.56 X-gal-negative cardiomyocytes/mm2 (p < 0.05). Using 5-ethynyl-2'-deoxyurinide (EdU) administration after MI, the percentages of EdU-positive CSP cells in the LIF-treated and control mice were 29.4 ± 2.7% and 10.6 ± 3.7%, respectively, which suggests that LIF influenced CSP proliferation. Moreover, LIF activated the Janus kinase (JAK)signal transducer and activator of transcription (STAT), mitogen-activated protein kinase/extracellular signal-regulated (MEK)extracellular signal-regulated kinase (ERK), and phosphatidylinositol 3-kinase (PI3K)-AKT pathways in CSPs in vivo and in vitro. The enhanced green fluorescent protein (EGFP)-bone marrow-chimeric CreLacZ mouse results indicated that LIF did not stimulate cardiogenesis via circulating bone marrow-derived cells during the 4 weeks following MI. Thus, LIF stimulates, in part, stem cell-derived cardiomyocyte regeneration by activating cardiac stem or precursor cells. This approach may represent a novel therapeutic strategy for cardiogenesis.

STAT transcription factors are induced by a number of growth factors and cytokines. Within minutes of induction, the STAT proteins are phosphorylated on tyrosine and serine residues and translocated to the nucleus, where they bind to their DNA targets. The leukemia inhibitory factor (LIF) mediates pleiotropic and sometimes opposite effects both in vivo and in cultured cells. It is known, for example, to prevent differentiation of embryonic stem (ES) cells in vitro. To get insights into LIF-regulated signaling in ES cells, we have analyzed protein-binding and transcriptional properties of STAT recognition sites in ES cells cultivated in the presence and in the absence of LIF. We have detected a specific LIF-regulated DNA-binding activity implicating the STAT3 protein. We show that STAT3 phosphorylation is essential for this LIF-dependent DNA-binding activity. The possibility that ERK2 or a closely related protein kinase, whose activity is modulated in a LIF-dependent manner, contributes to this phosphorylation is discussed. Finally, we show that the multimerized STAT3-binding DNA element confers LIF responsiveness to a minimal thymidine kinase promoter. This, together with our observation that overexpression of STAT3 dominant-negative mutants abrogates this LIF responsiveness, clearly indicates that STAT3 is involved in LIF-regulated transcriptional events in ES cells. Finally, stable expression of such a dominant negative mutant of STAT3 induces morphological differentiation of ES cells despite continuous LIF supply. Our results suggest that STAT3 is a critical target of the LIF signaling pathway, which maintains pluripotent cell proliferation.

Cell transplantation is a promising therapy for several muscle diseases, including Duchenne muscular dystrophy. Satellite cells are stem cells in skeletal muscle that provide an important cell source for transplantation therapy. However, culture of satellite cells in vitro causes them to lose their undifferentiated state, associated with reduced transplantation efficiency. It is therefore necessary to develop optimal culture conditions for maintaining the undifferentiated state of satellite cells.

The role of the IL6 family members in organ fibrosis, including renal interstitial fibrosis (TIF), has been widely explored. However, few studies have ever simultaneously examined them in the same cohort of patients. Besides, the role of leukemia inhibitory factor (LIF) in TIF remains unclear.

Expression of leukemia inhibitory factor (LIF) by a subset of Müller glia cells has recently been implicated in an endogenous survival response to photoreceptor injury in a model of inherited retinal degeneration. To investigate whether such a LIF-controlled survival pathway might be commonly induced upon photoreceptor injury independently of the nature of the toxic stimulus, we analyzed the role of LIF during light-induced retinal degeneration.

Implantation requires communication between a receptive endometrium and a healthy blastocyst. This maternal-embryonic crosstalk involves local mediators within the uterine microenvironment. We demonstrate that a secreted protein, prokineticin 1 (PROK1), is expressed in the receptive endometrium and during early pregnancy. PROK1 induces expression of leukemia inhibitory factor (LIF) in endometrial epithelial cells and first trimester decidua via a Gq-Ca(2+)-cSrc-mitogen-activated protein kinase kinase-mediated pathway. We show that human embryonic chorionic gonadotropin (hCG) induces sequential mRNA expression of PROK1 and LIF in an in vivo baboon model, in human endometrial epithelial cells, and in first-trimester decidua. We have used micro RNA constructs targeted to PROK1 to demonstrate that hCG-mediated LIF expression in the endometrium is dependent on prior induction of PROK1. Dual immunohistochemical analysis colocalized expression of the luteinizing hormone/chorionic gonadotropin receptor, PROK1, PROKR1, and LIF to the glandular epithelial cells of the first trimester decidual tissue. PROK1 enhances adhesion of trophoblast cells to fibronectin and laminin matrices, which are mediated predominantly via LIF induction. These data describe a novel signaling pathway mediating maternal-embryonic crosstalk, in which embryonic hCG via endometrial PROK1 may play a pivotal role in enhancing receptivity and maintaining early pregnancy.

The pro-myelinating effects of leukemia inhibitory factor (LIF) and other cytokines of the gp130 family, including oncostatin M (OSM) and ciliary neurotrophic factor (CNTF), have long been known, but controversial results have also been reported. We recently overexpressed erythropoietin receptor (EPOR) in rat central glia-4 (CG4) oligodendrocyte progenitor cells (OPCs) to study the mechanisms mediating the pro-myelinating effects of erythropoietin (EPO). In this study, we investigated the effect of co-treatment with EPO and LIF.

Leukemia inhibitory factor (LIF) is a multi-function cytokine. Its role in cancer is not well-understood. Recent studies including ours show that LIF is frequently overexpressed in many types of human tumors and promotes the progression and metastasis of tumors. However, the underlying mechanism of LIF's promoting effects on tumor progression and metastasis is poorly defined. Epithelial-mesenchymal transition (EMT) plays an important role in tumor metastasis. This study reports that LIF promotes EMT in human tumor cells. Overexpression of LIF promotes tumor cells to acquire mesenchymal features, including morphological changes of cells from epithelial-like to mesenchymal-like, increased expression levels of mesenchymal markers and decreased expression of epithelial markers. Knockdown of endogenous LIF reverses EMT in cancer cells. We further identified that LIF induces the expression of microRNA-21 (miR-21), which in turn mediates the promoting effect of LIF on EMT. LIF induces miR-21 expression through the activation of STAT3. Importantly, blocking miR-21 function greatly abolished the promoting effect of LIF on EMT and the migration ability of cancer cells. Taken together, results from this study identified an important function and a novel underlying mechanism of LIF in EMT and tumor metastasis.

LIF, a multifunctional cytokine, is frequently overexpressed in many types of solid tumors, including breast cancer, and plays an important role in promoting tumorigenesis. Currently, how LIF promotes tumorigenesis is not well-understood. Metabolic reprogramming is a hallmark of cancer cells and a key contributor to cancer progression. However, the role of LIF in cancer metabolic reprogramming is unclear. In this study, we found that LIF increases glucose uptake and drives glycolysis, contributing to breast tumorigenesis. Blocking glucose uptake largely abolishes the promoting effect of LIF on breast tumorigenesis. Mechanistically, LIF overexpression enhances glucose uptake via activating the AKT/GLUT1 axis to promote glycolysis. Blocking the AKT signaling by shRNA or its inhibitors greatly inhibits glycolysis driven by LIF and largely abolishes the promoting effect of LIF on breast tumorigenesis. These results demonstrate an important role of LIF overexpression in glucose metabolism reprogramming in breast cancers, which contributes to breast tumorigenesis. This study also reveals an important mechanism underlying metabolic reprogramming of breast cancers, and identifies LIF and its downstream signaling as potential therapeutic targets for breast cancers, especially those with LIF overexpression.

Respiratory syncytial virus (RSV) infects the lung epithelium where it stimulates the production of numerous host cytokines that are associated with disease burden and acute lung injury. Characterizing the host cytokine response to RSV infection, the regulation of host cytokines and the impact of neutralizing an RSV-inducible cytokine during infection were undertaken in this study.

Leukemia inhibitory factor (LIF) is a multifunctional cytokine. The present study aimed to determine the expression and effects of LIF on nucleus pulposus generation. Degenerated nucleus pulposus samples were obtained from animal models and patients with lumbar intervertebral disc herniation. Degradation scores of intervertebral discs were evaluated via magnetic resonance imaging (MRI) and histology, and the protein expression levels of LIF were detected. Furthermore, cultured primary human degenerated nucleus pulposus cells (DNPCs) were stimulated with various concentrations of recombinant human LIF protein (rhLIF), and aggrecan and collagen type II α1 (COL2α1) protein expression levels were detected by western blotting. In addition, aggrecan expression was determined by toluidine blue staining. The effects of rhLIF on proliferation and apoptosis of DNPCs were evaluated by Cell Counting Kit‑8 and flow cytometry, respectively. The results revealed that the degradation scores of intervertebral discs were significantly associated with modeling time, as determined by MRI and histology. In addition, the protein expression levels of LIF were initially increased in patients with lumbar disc herniation and in rabbit models, particularly in the 2‑week modeling group; however, its expression decreased with the progression of disc degeneration. Notably, LIF expression in each modeling group was higher than that in the control and 0 week modeling group. The in vitro study revealed that the protein expression levels of aggrecan and COL2α1 were significantly increased in response to rhLIF, in a dose‑dependent manner, and statistical differences were identified between the treatment groups and control group. The results of toluidine blue staining were consistent with this finding. Although rhLIF had no effect on proliferation, it inhibited apoptosis of DNPCs in a concentration‑dependent manner. In conclusion, LIF was upregulated during the process of intervertebral disc degeneration, and may promote the expression of extracellular matrix components. It may also be hypothesized that LIF acts as a potential protective factor by inhibiting apoptosis of DNPCs without affecting cell proliferation.

The consumption of large amounts of dietary fats can trigger an inflammatory response in the hypothalamus and contribute to the dysfunctional control of caloric intake and energy expenditure commonly present in obesity. The objective of this study was to identify chemokine-related transcripts that could be involved in the early stages of diet-induced hypothalamic inflammation.

Extramedullary hematopoiesis (EMH) expands hematopoietic capacity outside of the bone marrow in response to inflammatory conditions, including infections and cancer. Because of its inducible nature, EMH offers a unique opportunity to study the interaction between hematopoietic stem and progenitor cells (HSPCs) and their niche. In cancer patients, the spleen frequently serves as an EMH organ and provides myeloid cells that may worsen pathology. Here, we examined the relationship between HSPCs and their splenic niche in EMH in a mouse breast cancer model. We identify tumor produced IL-1α and leukemia inhibitory factor (LIF) acting on splenic HSPCs and splenic niche cells, respectively. IL-1α induced TNFα expression in splenic HSPCs, which then activated splenic niche activity, while LIF induced proliferation of splenic niche cells. IL-1α and LIF display cooperative effects in activating EMH and are both up-regulated in some human cancers. Together, these data expand avenues for developing niche-directed therapies and further exploring EMH accompanying inflammatory pathologies like cancer.

It has been demonstrated that leukemia inhibitory factor (LIF) induces epithelium apoptosis through Stat3 activation during mouse mammary gland involution. In contrast, it has been shown that this transcription factor is commonly activated in breast cancer cells, although what causes this effect remains unknown. Here we have tested the hypothesis that locally produced LIF can be responsible for Stat3 activation in mouse mammary tumors.

Progestogens including progesterone (P4) and levonorgestrel (LNG) are clinically used for multiple purposes such as contraception and infertility treatment. The effects of progestogens on the uterus remains to be elucidated. Here we examine the effect of excessive progestogen administration on embryo implantation focusing on the function of uterine leukemia inhibitory factor (LIF), a cytokine that is induced by estrogen and essential for embryo attachment. Treatment of wild-type (WT) female mice with vehicle (control), LNG at the dose of 300 μg/kg/day and P4 at the dose of 10 mg/day from day 1 to day 4 of pregnancy was conducted. LNG-treated and P4-treated mice showed embryo attachment failure on day 5 of pregnancy (The rate of mice with embryo attachment sites [%MAS], 11% and 13%, respectively), while all the control mice had normal attachment sites. Uterine LIF expression was significantly reduced in LNG-treated and P4-treated mice on day 4 evening. Administration of recombinant LIF (rLIF) at the dose of 24 μg/day on day 4 significantly rescued embryo attachment failure in LNG-treated and P4-treated mice (%MAS, 80% and 75%, respectively). Estradiol (E2) administration also rescued embryo attachment failure in LNG-treated mice (%MAS, 83%). Furthermore, excess P4 treatment before implantation decreased decidual P4 receptor (PGR) expression and induced decidualization defect apart from LIF downregulation. These findings indicate that progestogens cause embryo attachment inhibition through downregulation of uterine LIF expression and compromised decidualization through downregulation of PGR independently of LIF reduction. This study may contribute to a better understanding of contraceptive action of progestogens.