Leukemia Inhibitory Factor Gene Research Articles

Epigenetic alteration of uterine Leukemia Inhibitory Factor gene after glyphosate or a glyphosate-based herbicide exposure in rats

Glyphosate-based herbicides (GBHs) or its active ingredient, glyphosate (Gly), induce implantation failure in rats. We aimed to elucidate a mechanism of action of these compounds assessing the transcriptional and epigenetic status of the receptivity marker, leukemia inhibitory factor (Lif) gene. F0 rats were orally exposed to GBH or Gly at 3.8 or 3.9 mg Gly/kg/day, respectively, from gestational day (GD) 9 until weaning. F1 females were mated and uterine samples collected at GD5. Methylation-sensitive restriction enzymes (MSRE) sites and transcription factors were in silico predicted in regulatory regions of Lif gene. DNA methylation status and histone modifications (histone 3 and 4 acetylation (H3Ac and H4Ac) and H3 lysine-27-trimethylation (H3K27me3)) were assessed. GBH and Gly decreased Lif mRNA levels and caused DNA hypermethylation. GBH increased H3Ac levels, whereas Gly reduced them; both compounds enhanced H3K27me3 levels. Finally, both GBH and Gly induced similar epigenetic alterations in the regulatory regions of Lif.

Effect of the LIF gene on the cell cycle and apoptosis of ovarian granulosa cells in white Muscovy ducks

Leukaemia inhibitory factor (LIF), a member of the interleukin-6 (IL-6) family, is a multifunctional cytokine. The maturation-to-ovulation process of poultry follicles is determined by granulosa cell proliferation and differentiation. Granulosa cell apoptosis and degeneration lead to follicular atresia, which reduces the number of normally developing follicles and leads to a decrease in the poultry egg production rate, thus affecting the large-scale development of poultry breeding. In this study, the LIF gene overexpression vector pCDH-CMV-LIF and a siRNA that inhibits LIF gene expression were transfected into primary granulosa cells from white Muscovy duck ovaries for functional study. Compared with that in the control group, LIF gene expression was confirmed to be significantly decreased or increased in the transfection groups (P < 0.01). After LIF overexpression, the expression of the cell cycle-related genes CCND1, CDK-1 and PCNA was decreased (P < 0.05); apoptosis was promoted; the proapoptotic genes Bax and caspase-3 were significantly upregulated (P < 0.01); and the antiapoptotic gene Bcl-2 was significantly downregulated (P < 0.01). After LIF interference, the expression of the cell cycle-related genes CCND1, CCNE1, CDK-1 and PCNA and the antiapoptotic gene Bcl-2 significantly increased (P < 0.01), whereas the expression of the proapoptotic genes Bax, caspase-3 and caspase-9 significantly decreased (P < 0.01). In summary, the LIF gene is involved in regulating the biological function of ovarian granulosa cells in white Muscovy ducks. LIF gene expression promotes granulosa cell apoptosis and inhibits cell cycle progression. These experimental results provide insights into the follicular development mechanism of white Muscovy ducks.

Identification and genetic validation of leukemia inhibitory factor super-enhancers in acute respiratory distress syndrome and lung cancer.

Super-enhancers play prominent roles in driving robust pathological gene expression, but they are hidden in human genome at noncoding regions, making them difficult to explore. Leukemia inhibitory factor (LIF) is a multifunctional cytokine crucially involved in acute respiratory distress syndrome (ARDS) and lung cancer progression. However, the mechanisms governing LIF regulation in disease contexts remain largely unexplored. In this study, we observed elevated levels of LIF in the bronchoalveolar lavage fluid (BALF) of patients with sepsis-related ARDS compared to those with nonsepsis-related ARDS. Furthermore, both basal and LPS-induced LIF expression were under the control of super-enhancers. Through analysis of H3K27Ac ChIP-seq data, we pinpointed three potential super-enhancers (LIF-SE1, LIF-SE2, and LIF-SE3) located proximal to the LIF gene in cells. Notably, genetic deletion of any of these three super-enhancers using CRISPR-Cas9 technology led to a significant reduction in LIF expression. Moreover, in cells lacking these super-enhancers, both cell growth and invasion capabilities were substantially impaired. Our findings highlight the critical role of three specific super-enhancers in regulating LIF expression and offer new insights into the transcriptional regulation of LIF in ARDS and lung cancer.

Determination of the Frequency of BCL-2 Polymorphisms (c.-717C>A and c.*2364G>A) and LIF Polymorphism (c.*1414T>G) in Patients with Congenital Anomalies of the Kidney and Urinary Tract

Introduction: Congenital anomalies of the kidney and urinary tract (CAKUT) are characterized by several malformations. Its prevalence is 0.3–0.6% in live births. The B-cell lymphoma (BCL-2) gene regulates apoptosis, and the Leukemia Inhibitory Factor (LIF) gene plays a role in many biological processes, such as blastocyst growth and uterine preparation for implantation. In this study, two single nucleotide polymorphisms (SNPs) of the BCL-2 gene (rs2279115 and rs4987856) and one SNP of the LIF gene (rs929271) were investigated in CAKUT patients for the first time. Methods: Hundred and twenty-nine CAKUT patients and 105 controls were enrolled in this study. We used polymerase chain reaction-restriction fragment length polymorphism for rs2279115 and rs929271 and SNaPshot for rs4987856. The χ2 test was used to compare discrete variables, and the independent sample t test was used to compare continuous variables. Results: The allele frequencies for the rs2279115 and rs4987856 polymorphisms of BCL-2 and the rs929271 polymorphism of LIF were not significantly different between the patient and control groups (p = 0.162, p = 0.053, p = 0.635, respectively). However, the co-segregation analysis revealed a significant difference in the distribution of allele frequencies between the patient and control groups for two genetic variations: LIF rs929271 SNP and BCL-2 rs4987856 SNP (p = 0.034). The relative odds ratio was 2.444 (95% Confidence Interval (CI) 1.054–5.671). Conclusion: This study, which is the first time in the literature, showed that changes in BCL-2 and LIF genes are associated with CAKUT disease.

Engineered exosome as a biological nanoplatform for drug delivery of Rosmarinic acid to improve implantation in mice with induced endometritis

Endometritis is an inflammatory and histopathologic disease in uterine tissues that interferes with the proper decidualization and implantation of the embryo. In this study, rosmarinic acid (RA) is used as an anti-inflammatory agent that encapsulates in exosomes and is used to attenuate lipopolysaccharide (LPS)-induced endometritis and improve implantation. For this purpose, exosomes were loaded with RA and then administrated into the animal groups, including RA, exosome, RA plus exosome (RA + Exo), and RA-loaded exosomes (RALExo) groups. The concentrations of RA or exosomes used in this study were 10 mg/kg, and the compounds were injected into the uterine horn 24 h following the induction of endometritis. Upon the presence of inflammation detected by the histopathological method, the most proper groups were mated with male mice. The effect of the treatment group on the implantation rate, progesterone levels, and gene expressions were assessed by Chicago Blue staining, enzyme-linked immunosorbent assay (ELISA), and Quantitative PCR (qPCR), respectively. Results showed RALExo10 and RA10 + Exo10 groups improved pathological alterations, enhanced progesterone levels, increased implantation rate, as well as heightened expression levels of Leukemia inhibitory factor (LIF) and Mucin-16 (MUC-16) genes. Besides, the expression levels of inflammatory cytokines, including Transforming growth factor-β (TGF-β), Interlukine-10 (IL-10), Interlukine-15 (IL-15), and Interlukine-18 (IL-18), were regulated. Our findings indicated that the expression of LIF, Muc-16 genes as well as IL-18, were significantly correlated with serum progesterone concentrations and the implantation rate in the treatment groups. The RALExo10 and RA10 + Exo10 groups showed ameliorated implantation rates in experimental groups.

Possible Role of Leukemia Inhibitory Factor and Inflammatory Cytokines in The Recurrent Spontaneous Abortion: A Case-Control Study.

Despite of long-lasting efforts, in more than 50% of cases, the etiology of recurrent spontaneous abortion (RSA) remains unknown. Leukemia inhibitory factor (LIF) has an essential role in the reproductive process, such as modulating inflammatory responses. This study aimed to evaluate the relationship between the LIF gene expression as well as serum levels of inflammatory cytokines and occurrence of RSA in infertile women with a history of RSA. In this case-control study, the relative gene expression levels of LIF, concentrations of tumor necrosis factor-alpha (TNF-α), and interleukin (IL)-17 were measured in peripheral blood and serum of women with a history of RSA (N=40) compared with non-pregnant and fertile women as the control group (N=40) using quantitative real-time polymerase chain reaction and the enzyme-linked immunosorbent assay, respectively. The mean age of patients and controls was 30.1 ± 4.28 and 30.03 ± 4.23, respectively. Patients had a history of at least 2 and at most 6 abortions. The mRNA levels of LIF were significantly lower in the women with RSA in comparison with the healthy participant (P=0.003). Regarding cytokine levels, no significant difference was seen between the two groups (P≥0.05). There was no correlation - between the LIF mRNA levels and TNF-α and IL-17 serum concentrations. The U-Mann-Whitney test and the Pearson correlation coefficient were applied to comparison variables between groups as well as a correlation between LIF mRNA and cytokine levels in serum. Despite a significant reduction in the LIF gene mRNA level in patients with RSA, it was not associated with increases in inflammatory cytokines. Dysfunction in the production of LIF protein may be involved in the onset of RSA disorder.

Association between Leukemia Inhibitory Factor Gene Polymorphism and Clinical Outcomes among Young Women with Poor Ovarian Response to Assisted Reproductive Technology.

Does the presence of single-nucleotide polymorphisms (SNPs) in the leukemia inhibitory factor (LIF) gene affect ovarian response in infertile young women? This was a case-control study recruiting 1744 infertile women between January 2014 to December 2015. The 1084 eligible patients were stratified into four groups using the POSEIDON criteria. The gonadotropin-releasing hormone receptor (GnRHR), follicle-stimulating hormone receptor (FSHR), anti-Müllerian hormone (AMH), and LIF SNP genotypes were compared among the groups. The distributions of LIF and FSHR among younger and older patients were compared. Clinical outcomes were also compared. The four groups of poor responders had different distributions of SNP in LIF. The prevalence of LIF genotypes among young poor ovarian responders differed from those of normal responders. Genetic model analyses in infertile young women revealed that the TG or GG genotype in the LIF resulted in fewer oocytes retrieved and fewer mature oocytes relative to the TT genotypes. In older women, the FSHR SNP genotype contributed to fewer numbers of mature oocytes. LIF and FSHR SNP genotypes were associated with a statistically significant reduction in ovarian response to controlled ovarian hyperstimulation in younger and older women with an adequate ovarian reserve, respectively.

Elevated expression of LIF predicts a poor prognosis and promotes cell migration and invasion of clear cell renal cell carcinoma.

BackgroundRenal cell carcinoma (RCC) is the seventh most common cancer in humans, of which clear cell renal cell carcinoma (ccRCC) accounts for the majority. Recently, although there have been significant breakthroughs in the treatment of ccRCC, the prognosis of targeted therapy is still poor. Leukemia inhibitory factor (LIF) is a pleiotropic protein, which is overexpressed in many cancers and plays a carcinogenic role. In this study, we explored the expression and potential role of LIF in ccRCC.MethodsThe expression levels and prognostic effects of the LIF gene in ccRCC were detected using TCGA, GEO, ICGC, and ArrayExpress databases. The function of LIF in ccRCC was investigated using a series of cell function approaches. LIF-related genes were identified by weighted gene correlation network analysis (WGCNA). GO and KEGG analyses were performed subsequently. Cox univariate and LASSO analyses were used to develop risk signatures based on LIF-related genes, and the prognostic model was validated in the ICGC and E-MTAB-1980 databases. Then, a nomogram model was constructed for survival prediction and validation of ccRCC patients. To further explore the drug sensitivity between LIF-related genes, we also conducted a drug sensitivity analysis based on the GDSC database.ResultsThe mRNA and protein expression levels of LIF were significantly increased in ccRCC patients. In addition, a high expression of LIF has a poor prognostic effect in ccRCC patients. LIF knockdown can inhibit the migration and invasion of ccRCC cells. By using WGCNA, 97 LIF-related genes in ccRCC were identified. Next, a prognostic risk prediction model including eight LIF-related genes (TOB2, MEPCE, LIF, RGS2, RND3, KLF6, RRP12, and SOCS3) was developed and validated. Survival analysis and ROC curve analysis indicated that the eight LIF-related-gene predictive model had good performance in evaluating patients’ prognosis in different subgroups of ccRCC.ConclusionOur study revealed that LIF plays a carcinogenic role in ccRCC. In addition, we firstly integrated multiple LIF-related genes to set up a risk-predictive model. The model could accurately predict the prognosis of ccRCC, which offers clinical implications for risk stratification, drug screening, and therapeutic decision.

P-327 The impact of chronic endometritis disease on endometrial receptivity gene expression in women with repeated implantation failure

Abstract Study question Does chronic endometritis (CE) have an impact on the expression of the genes involved in the embryo implantation? Summary answer The mRNA expression of genes involved in embryo implantation (HOXA10, HOXA11, BTEB1 and LIF) does not change in patients with and without CE. What is known already CE is an inflammatory disorder of the endometrium with a detrimental effect on embryo implantation and its prevalence in women with repeated implantation failure (RIF) is up to 30%. During the implantation window, the expression of some genes plays a key role in determining the success of embryo implantation. The homeobox genes HOXA10, HOXA11 and the basic transcriptional element binding protein 1 (BTEB1) gene are essential for endometrial cells growth regulation and for embryonic development and the leukaemia inhibitory factor (LIF) has a role in the embryo–endometrium interaction. Study design, size, duration A single centre-prospective case-control study was conducted at the Women’s and Children’s Health Department at Padua University, on infertile women with RIF from June 2020 to November 2021. 29 women with RIF were prospectively enrolled. Participants/materials, setting, methods An endometrial biopsy using a Novak curette was performed for each patient and the obtained tissue samples were divided in two aliquots, one for immunohistochemistry for histological examination (endometrial dating and CE diagnosis) and one for RNA extraction and gene expression analysis that has been performed using RT-PCR. Sample of patients with and without CE were compared using non-parametric Mann–Whitney U-test. A p-value &amp;lt;.05 was considered as statistically significant. Main results and the role of chance Enrolled women were divided in two groups according to the histological diagnosis of CE: 13 patients with CE, 16 without CE. In all samples, appropriate histological dating (WOI) was evaluated according to Noyes criteria. After the comparison of all investigated genes (HOXA10, HOXA11, BTEB1 and LIF), no significant difference in mRNA expression was detected between women with and without CE (p value &amp;gt;.05). In the literature, defective endometrial expression of HOXA10, HOXA11 and LIF genes has been associated with abnormal implantation and the reduced expression of BTEB1 gene results in subfertility and progesterone resistance. Although the negative role of CE in altering embryo implantation is known, our results suggest that the CE has no effect on the expression of HOXA10, HOXA11, BTEB1 and LIF genes. Probably the detrimental effect of CE on embryo implantation does not act through HOXA10, HOXA11, BTEB1 and LIF gene expression. Limitations, reasons for caution This is a non-randomized observational study with a limited number of patients. Further studies are needed to confirm our data with immunohistochemistry evaluation to define the protein expression levels of the investigated genes. Wider implications of the findings Understanding the pathogenic mechanism of CE on endometrial receptivity is crucial for identifying markers that best correlate with possible implantation failure and for identifying the appropriate therapy to treat the disease and restore the embryo implantation capacity. Trial registration number not applicable

Altered expression of leukemia inhibitory factor (LIF), LIFR, gp130, and IL11 in the embryo implantation site of rat model with prenatal androgen-induced polycystic ovary syndrome

Polycystic ovary syndrome (PCOS) is a complex endocrine disorder that represents infertility in many reproductive-age women. Reduced implantation of blastocyst was proposed as an etiology for infertility in this syndrome. In this regard, many candidate genes such as leukemia inhibitory factor (LIF), LIF receptor (LIFR), glycoprotein 130 (gp130), and interleukin 11 (IL11) were proposed to be disrupted. Investigation of these genes is not ethically approved in pregnant women with PCOS. In this study, we aimed to compare the expression of LIF, LIFR, gp130, and IL11 before and during different gestational days in uterine tissues of prenatally-androgenized rat models of PCOS with control rats. The rat model of polycystic ovary syndrome was created by the injection of testosterone during prenatal life. RNA extraction and cDNA synthesis from uterine tissues were performed in both prenatal induced PCOS and control rats. Expression of LIF, LIFR, gp130, and IL11 genes was compared before pregnancy (GD0) and during pregnancy on GD0.5, GD4.5, GD5.5, and GD8.5 between two study groups (n = 6 each group) using SYBR Green real-time PCR. The expression of the LIF mRNAs significantly decreased on GD4.5, 5.5, and 8.5 in the PCOS rats compared to the controls (P-values: 0.0483, 0.0152, and 0.0043). Additionally, decreased expression of LIFR and gp130 was observed on GD0.5 to 8.5 in PCOS rats compared to controls (P-values: 0.022, 0.0480, 0.0043, 0.0022 for LIFR and 0.0189, 0.0022, 0.0087, 0.0022 for gp130). Moreover, IL-11 mRNA levels decreased in the PCOS group compared to their controls both before (P-value:0.0362) and during the gestational period (P-values:0.0085, 0.0043, 0.0389, 0.0087). Reduced expression of LIF, LIFR, gp130, and IL11 in the rats with PCOS indicates a possible disruption in the implantation and decidualization stages in this syndrome.

Endometrial injury concurrent with hysteroscopy increases the expression of Leukaemia inhibitory factor: a preliminary study

ObjectiveIt is not known by which mechanism endometrial injury increases pregnancy rates. Leukaemia inhibitory factor (LIF) is a cytokine involved in wound healing and implantation. The aim of this study was to determine the change in endometrial LIF mRNA expression before and after mechanical injury during hysteroscopy.MethodsForty patients with a history of two or more unsuccessful implantations who decided to undergo hysteroscopy in the proliferative phase were divided into two equal groups: one with endometrial injury (scratching group) and the other with noninjury (control group). Endometrial sampling was conducted before injury on the patients in the scratching group, and then injury was performed with monopolar needle forceps. Only diagnostic hysteroscopy was performed on the patients in the control group. Endometrial tissues were collected using a Pipelle catheter between Days 20 and 23 of the mid-luteal phase of the next cycles in both the scratching and control groups. Endometrial LIF mRNA expression was evaluated with the use of reverse-transcription polymerase chain reactions.ResultsRelative changes in mRNA expression levels of the LIF gene in endometrial samples taken before and after injury were calculated using the 2-ΔΔCt method, and the fold changes obtained were compared between and within the groups. Compared with preinjury values, an 11.1-fold increase was found in postinjury LIF mRNA expression in patients with monopolar forceps injury (p < 0.001). There was a 3.9-fold significant increase in postinjury LIF mRNA levels compared with those in the control group (p < 0.02).ConclusionsThe fertility-promoting effect of hysteroscopy-guided mechanical endometrial injury may be mediated by LIF mRNA.

The enhancer rare germline variation rs548071605 contributes to lung cancer development.

Rare germline variations contribute to the missing heritability of human complex diseases including cancers. Given their very low frequency, discovering and testing disease-causing rare germline variations remains challenging. The tag-single nucleotide polymorphism rs17728461 in 22q12.2 is highly associated with lung cancer risk. Here, we identified a functional rare germline variation rs548071605 (A>G) in a p65-responsive enhancer located within 22q12.2. The enhancer significantly promoted lung cancer cell proliferation in vitro and in a xenograft mouse model by upregulating the leukemia inhibitory factor (LIF) gene via the formation of a chromatin loop. Differential expression of LIF and its significant correlation with first progression survival time of patients further supported the lung cancer-driving effects of the 22q-Enh enhancer. Importantly, the rare variation was harbored in the p65 binding sequence and dramatically increased the enhancer activity by increasing responsiveness of the enhancer to p65 and B-cell lymphoma 3 protein, an oncoprotein that assisted the p65 binding. Our study revealed a regulatory rare germline variation with a potential lung cancer-driving role in the 22q12.2 risk region, providing intriguing clues for investigating the "missing heritability" of cancers, and also offered a useful experimental model for identifying causal rare variations.

The effect of cytokine leukemia-inhibitory factor (LIF) and interleukin-11 (IL-11) gene expression on the primary infertility related to polycystic ovary syndrome, Tubal factor, and Unexplained infertility in Turkish women

BackgroundSuccessful implantation of blastocysts is indeed an important path in mammalian reproduction that is governed by a complicated web of cytokines interactions. Leukemia inhibitory factor (LIF) and interleukin-11 (IL-11) part of the interleukin (IL)-6 groups are cytokines that are needed for effective implantation and prevent infertility symptoms. This study aimed to determine the expression level (LIF, IL-11) genes in patients with primary infertility related to polycystic ovary syndrome (PCOS), tubal factor infertility (TFI), and unexplained infertility (UI).ResultsIn this study, 75 infertility women and 40 controls were involved. The expressions of LIF and IL-11 genes were evaluated by quantitative real-time polymerase chain reaction qRT–PCR Light Cycler in patients and healthy controls. PCOS, TFI, and UI groups showed promising results regarding LIF gene, which appeared at very small levels compared to the control (p < 0.0001). Regarding IL-11, the two groups TFI and UI were significantly linked to the lower level of gene expression, while the PCOS group has no significant difference when it is compared to the control group (p < 0.0001, < 0.05, 0.19), respectively.ConclusionThe current findings show that low levels of LIF and IL-11 gene expression are linked to various primary infertility conditions, including PCOS, tubal factor, and unexplained infertility since they play a fundamental role in embryo implantation.

Protective effect of LIF-huMSCs on the retina of diabetic model rats.

To investigate the protective effect of human umbilical cord mesenchymal stem cells (hUCMSCs) modified by the LIF gene on the retinal function of diabetic model rats and preliminarily explore the possible mechanism. A stably transfected cell line of hUCMSCs overexpressing leukemia inhibitory factor (LIF) was constructed. Overexpression was verified by fluorescent quantitative polymerase chain reaction (qPCR). Forty-eight adult Sprague-Dawley rats were randomly divided into a normal control group (group A), streptozotocin-induced diabetic control group (group B), diabetic rats at 3mo injected with empty vector-transfected hUCMSCs (group C) or injected with LIF-hUCMSCs (group D). Four weeks after the intravitreal injection, analyses in all groups included retinal function using flash electroretinogram (F-ERG), retinal blood vessel examination of retinal flat mounts perfused with fluorescein isothiocyanate-dextran (FITC-dextran), and retinal structure examination of sections using hematoxylin and eosin staining. Expression levels of adiponectin (APN), high-sensitivity C-reactive protein (hs-CRP), and neurotrophin-4 (NT-4) in each group was detected using immunohistochemistry, PCR, Western blotting, and ELISA, respectively. A stable transgenic cell line of LIF-hUCMSCs was constructed. F-ERG and FITC-dextran examinations revealed no abnormalities of retinal structure and function in group A, severe damage of the retinal blood vessels and function in group B, and improved retinal structure and function in group C and especially group D. qPCR, ELISA, and Western blot analyses revealed progressively higher APN and NT-4 expression levels in groups B, C, and D than in group A. hs-CRP expression was significantly higher in group B than in groups A, C, and D, and was significantly higher in group C than in group D (P<0.05). LIF-hUCMSCs protect the retina of diabetic rats by upregulating APN and NT-4 expression and downregulating hs-CRP expression in the retina.

Investigation of Polymorphisms in the Upstream Sequence of LIF and LIFR Genes in the Infertile Women

: During pregnancy, the embryo implantation stage is a highly dynamic and molecularly controlled phenomenon. Several genes are involved in the implantation process, among which the leukemia inhibitory factor (LIF) is a marker of implantation. LIF is a multi-functional cytokine located on chromosome 22. The expression of this gene is increased in the middle of the secretion phase from the sexual cycle, and any defect in its expression will cause the implantation failure. LIF receptor or LIFR gene, as the LIF receptor, consists of two membrane proteins called LIFR and GP130. LIFR acts as a signal receptor for LIF in a low-affinity level. In this study, we focused on the screening of polymorphisms in the promoter region of the LIF and LIFR genes in the infertile women using the polymerase chain reaction single-strand conformation polymorphism (PCR-SSCP) method. In this study, blood samples were collected from 100 women with primary and secondary infertility and 50 healthy women as a control group. Extraction of DNA was done by the phenol-chloroform method, and in the next step, using specific primers for upstream regions of the LIF and LIFR genes, target sequences were amplified and analyzed by the SSCP method. Finally, PCR products with different configurations were selected for sequencing analysis. The results showed two polymorphisms in the upstream region of LIF and LIFR genes of two women, but there were no genetic changes in the control group. The present study was the first in this field, and the results indicated the importance of examining such genes in infertility with an unknown cause.

Effect of leukemia inhibitory factor polymorphism on litter size traits in Thai commercial pig breeds

Leukemia inhibitory factor (LIF) is a crucial candidate gene that impacts on implantation process. In this study, the effects of the porcine LIF polymorphism on litter size traits were elucidated in Thai commercial pig populations. Genotyping of three single nucleotide polymorphisms (SNPs) of the porcine LIF gene was detected in coding and 3ˊ-untranslated regions. The porcine LIF c.*24C&gt;T was segregating in Large White, Landrace, and Large White × Landrace (LW × LR) crossbred sows. No polymorphisms at two non-synonymous SNPs loci (LIF c.28C&gt;A and LIF c.161A&gt;G) were found in this study. The porcine LIF c.*24C&gt;T was significantly associated with the total number born (TNB), the number born alive (NBA), and the number of piglets weaned alive (NWA) traits in Large White and Landrace sows. Moreover, the porcine LIF c.*24C&gt;T was associated with the NBA and NWA traits in LW × LR crossbred sows. The favorable LIF c.*24C allele was positively correlated with the litter size traits. These findings indicated that the polymorphism of the porcine LIF gene was associated with litter size traits and confirms the significance of porcine LIF as a candidate gene for litter size traits in pig breeding. Thus, the porcine LIF gene could be used for improving prolific traits in these Thai commercial pig populations.

Epigenetic Profiling Identifies LIF as a Super-enhancer-Controlled Regulator of Stem Cell-like Properties in Osteosarcoma.

Osteosarcoma is an aggressive malignancy with poor prognosis. Super-enhancers (SE) have been highlighted as critical oncogenic elements required for maintaining the cancer cell characteristics. However, the regulatory role of SEs in osteosarcoma properties has not yet been elucidated. In the current study, we found that osteosarcoma cells and clinical specimens shared a significant fraction of SEs. Moreover, leukemia-inhibitory factor (LIF) was identified as an essential factor under the control of osteosarcoma-specific SE. The expression of LIF was positively correlated with the stem cell core factor genes in osteosarcoma. Furthermore, LIF recombinant protein-treated osteosarcoma cells displayed enhanced stem cell-like characteristics, such as increased sphere-forming potential, stimulated self-renewal, upregulated metastasis ability, and increased stemness-related gene expression. Notably, the histone 3 lysine 27 tri-methylation (H3K27me3) demethylase UTX was found as a key activator of LIF transcription in osteosarcoma. The UTX inhibitor, GSK-J4, induced H3K27me3 accumulation and impaired histone 3 lysine 27 acetylation (H3K27ac) at LIF gene locus, leading to LIF signaling pathway inhibition. GSK-J4 treatment resulted in profound defects in stem cell-like characteristics and stemness-related gene activation in osteosarcoma by modulating the H3K27ac of NOTCH1 signaling pathway gene loci. The NOTCH1 inhibitor Crenigacestat (TargetMol, T3633) repressed LIF-mediated activation of the stemness-related genes in osteosarcoma patient-derived primary tissues. IMPLICATIONS: This study reveals osteosarcoma SE profiles and uncovers a distinct tumor-stemness epigenetic regulatory mechanism in which an osteosarcoma-specific SE-mediated factor, LIF, promotes osteosarcoma stemness gene activation via NOTCH1 signaling pathway.

Neural Stem Cells Transfected with Leukemia Inhibitory Factor Promote Neuroprotection in a Rat Model of Cerebral Ischemia.

Leukemia inhibitory factor (LIF) contributes to the neuroprotection by neural stem cells (NSCs) after ischemic stroke. Our aim was to explore whether LIF-transfected NSCs (LIF-NSCs) can ameliorate brain injury and promote neuroprotection in a rat model of cerebral ischemia. To accomplish this goal, we transfected NSCs with a lentivirus carrying the LIF gene to stably overexpress LIF. The LIF-NSCs reduced caspase 3 activation under conditions of oxygen-glucose deprivation in vitro. Transient cerebral ischemia was induced in rats by 2h of middle cerebral artery occlusion (MCAo), and LIF-NSCs were intravenously injected at 6h post-ischemia. LIF-NSC treatment reduced the infarction volume and improved neurological recovery. Moreover, LIF-NSCs improved glial cell regeneration and ameliorated white matter injury in the MCAo rats. The NSCs acted as carriers and increased the expression of LIF in the lesions to protect against cerebral infarction, suggesting that LIF-NSCs could be a potential treatment for cerebral infarction.

Effects of rs929271 SNP in Leukemia Inhibitory Factor Gene on Recurrent Pregnancy Loss, Placental Location and Fetal Gender

: Leukemia inhibitory factor (LIF) has an essential role in embryo implantation and placentation. The purpose of this study was to investigate the association of rs929271 SNP on LIF gene with recurrent pregnancy loss (RPL), placental location, and fetal gender in Iranian Azeri women. A total of 300 Azeri women (150 women with at least two recurrent miscarriages and 150 women with at least two healthy deliveries, as the control group) were genotyped for the 3’UTR region T>G SNP on LIF gene (rs929271) by real-time polymerase chain reaction (PCR) method. Placental location and fetal gender were determined by two-dimensional sonography. The analysis showed that female embryos were more likely to abort than males (OR = 1.806, CI = 1.275 to 2.557, P value = 0.00083). The highest odds percentage ratio (OPR) of abortion were in women with fundal (OPR = 1.817) and anterior locations (OPR = 1.483); and the lowest in lower-lying placental locations (OPR = 0.448) (P = 0.0284). In this study, the frequency of T and G alleles were 0.99 and 0.01, respectively, and only two TT and TG genotypes were identified. Women with TG genotype had significantly more recurrent pregnancy loss than TT genotypes (P = 0.0133). The women with TG genotype compared with the TT genotypes were more likely to have low lying (OPR = 2.614) and anterior (OPR = 1.641) placental position. Also, the lowest OPR (0.231) for placental position of TG versus the TT genotype was posterior location (P = 0.0464). The allele G caused more (about 1.9 folds) female embryos than T allele, yet the difference was not significant (P = 0.34). In general, there are indications that the effect of this polymorphism on RPL is repeated due to its effect on the placental location and fetal gender. However, to confirm these results, there is a need to repeat this study in populations with more frequent G allele as well as other polymorphisms on the LIF gene.

Establishment of STO Cell Lines Expressing Green Fluorescent Protein and Mouse Leukemia Inhibitory Factor

To establish the STO cell lines expressing green fluorescent protein (GFP) and mouse leukemia inhibitory factor (LIF) , and try to culture the mouse embryonic stem cells (mESCs) by using the established STO-GFP-mLIF cells as the feeder layer. The lentiviral particles containing GFP and mLIF and puromycin-resistance gene were constructed and transduced into STO cell lines. The cell lines stably expressing GFP and mLIF genes were screened out. The expression level of the inserted exogenous LIF gene was tested by Western blot and ELISA. The STO-GFP-mLIF cells were treated with different concentrations of mitomycin C (5, 10, 15, 20 µg/ml) for different time (1.5, 2.5, 3, 3.5 hours) to prepare feeder layers and the cell proliferation level on feeder layer was observed. Mouse embryonic stem cells were cultured on mitomycin C-treated feeder layer and the growth of cell colonies was observed. The expression level of LIF protein in STO-GFP-mLIF cells was up-regulated, as compared with STO cells (P＜0.05). It was confirmed that the optimal concentration and time for inhibiting the proliferetion of STO-GFP-mLIF cells by mitomycin C were 10 µg/ml and 3 hours respectively. The observation also found that the embryonic stem cells could develop into typic "birdnest" shaped stem cell colony on mitomycin C-treated feeder layer. The stable STO cell lines effectively expressing green fluorescent protein and mouse leukemia inhibitory factor have been established successfully, which can maintain the undifferentiated state of mouse embryonic stem cells.