Expression of leukemia inhibitory factor in craniopharyngioma

It has recently been reported that overexpression of leukemia inhibitory factor (LIF) in mice transgenic for LIF causes invagination of the anterior wall of Rathke’s pouch leading to the formation of cysts lined by LIF immunoreactive epithelial cells. Strong immunoreactivity was also found in human Rathke’s cleft cysts. Because such cysts and craniopharyngiomas share a common histogenesis, we raised the question of whether LIF is also expressed in craniopharyngioma. Fourteen histologically verified craniopharyngiomas of adamantinomatous type were examined for LIF immunoreactivity using the streptavidin-biotin-peroxidase complex method. Rabbit-anti-LIF antibody dilution 1:40) was applied to tissues having undergone antigen retrieval (microwaving in citrate buffer at pH 6). For positive control, nontumorous pituitary tissues were used. Primary antibody substituted with phosphate-buffered saline served as a negative control. By immunocytochemistry, the epithelial cells of all 14 craniopharyngiomas were LIF immunoreactive, showing varying degrees of staining intensity. In comparison, the connective tissue components of the tumors were immunonegative. Our study provides evidence that LIF is expressed in the epithelial cells of craniopharyngioma. Further investigation is required to elucidate the possible role of LIF in the development and progression of craniopharyngiomas.     

The impact of leukemia inhibitory factor in uterine flushing on the reproductive potential of infertile women--a prospective study

PROBLEM: To determine the value of leukemia inhibitory factor (LIF) assessment for predicting the reproductive outcome. METHOD OF STUDY: Two phase study. Phase I: assessment of LIF in uterine flushing. Phase II: 1,5 years after examining the last patient, a questionnaire was sent to all participants of the phase I. Phase I: Uterine flushing and endometrial samples were collected during implantation window from infertile patients with stage I/II endometriosis (n = 14), patients with idiopathic infertility (n = 27), luteal phase deficiency (n = 13), and fertile control (n = 21). LIF was assessed in uterine flushings in all patients by ELISA. In endometrium, semiquantitative RT-PCR was performed for LIF mRNA expression. Phase II: questionnaire has been sent to all infertile women taking part in the first phase of the experiment, regarding their reproductive outcome. RESULTS: 65.4% patients who had returned the questionnaire did get pregnant. LIF concentration at a cut-off point of 2.31 pg/ml had a 95.7% sensitivity and 81.8% specificity in predicting the reproductive outcome. CONCLUSION: This prospective study for the first time in literature indicates that the LIF assessment can be used as a predictor of reproductive success.     

The unsolved enigmas of leukemia inhibitory factor

Leukemia inhibitory factor (LIF) is a polyfunctional glycoprotein cytokine whose inducible production can occur in many, perhaps all, tissues. LIF acts on responding cells by binding to a heterodimeric membrane receptor composed of a low-affinity LIF-specific receptor and the gp130 receptor chain also used as the receptor for interleukin-6, oncostatin M, cardiotrophin-1, and ciliary neurotrophic factor. LIF is essential for blastocyst implantation and the normal development of hippocampal and olfactory receptor neurons. LIF is used extensively in experimental biology because of its key ability to induce embryonic stem cells to retain their totipotentiality. LIF has a wide array of actions, including acting as a stimulus for platelet formation, proliferation of some hematopoietic cells, bone formation, adipocyte lipid transport, adrenocorticotropic hormone production, neuronal survival and formation, muscle satellite cell proliferation, and acute phase production by hepatocytes. Unwanted actions of LIF can be minimized by circulating soluble LIF receptors and by intracellular suppression by suppressors of cytokine-signaling family members. However, the outstanding problems remain of how the induction of LIF is mediated in response to demands from such a heterogeneity of target tissues and why it makes design sense to use LIF in the regulation of such a diverse and unrelated series of biological processes.     

Endogenous leukemia inhibitory factor attenuates endotoxin response

Leukemia inhibitory factor (LIF) is induced in inflammation and likely plays a regulatory role. Using LIF-deficient mice (LIF-/-), we report here that endogenous LIF has a protective role in endotoxic shock and host defence. LIF-/- mice have heightened sensitivity to LPS in a LPS/D-galactosamine (D-Gal) sensitization model compared to wild-type mice (LIF+/+), enhanced thrombocytopenia and leukopenia, with increased hepatic necrosis, neutrophil sequestration in the lung and accelerated mortality. These findings correlated with 10-fold higher tumour necrosis factor-alpha (TNFalpha) and interleukin-6 (IL-6) serum levels and reduced IL-10 production in LIF-/- mice in response to LPS. Therefore, endogenous LIF attenuates the endotoxic shock response, enhances the expression of basal acute phase proteins and IL-10 production, which downregulates TNFalpha synthesis and release and thereby confers partial protection to endotoxemia.     

正常女性子宫内膜LIF基因表达研究

本文采用RT－PCR技术对32例正常女性分泌期子宫内膜LIF基因表达进地了研究,结果在32例女性这一时期子宫内膜组织中均检测到有强烈的LIF基因表达。这提示在人类非妊娠分泌期子宫内膜组织中存在LIF基因表达LIF在胚泡着床中可能起着十分重要的作用。     

神经生长因子和白血病抑制因子上调哮喘大鼠脾淋巴细胞IL-4、IL-5的表达

目的:探讨神经生长因子(NGF)、白血病抑制因子(LIF)对哮喘大鼠脾淋巴细胞IL-4、IL-5表达的影响。方法:16只大鼠随机分为对照组(A组)8只,哮喘组(B组)8只,采用卵蛋白和氢氧化铝腹腔内注射致敏,2周后给予1%卵蛋白雾化吸入激发哮喘。(1%卵蛋白/生理盐水)雾化吸入2周后,24h内体外分离培养对照组大鼠和哮喘组大鼠脾淋巴细胞;通过RT-PCR半定量法和ELISA法测定2组大鼠脾淋巴细胞表达Th2细胞因子IL-4、IL-5mRNA转录水平和分泌蛋白的基础水平,观察外源性NGF和LIF体外干预后淋巴细胞IL-4和IL-5表达随干预浓度的变化。结果:(1)哮喘组IL-4、IL-5mRNA表达及蛋白水平明显高于对照组(P0.05);(2)在外源性不同浓度的NGF、LIF干预下,IL-4、IL-5mRNA和蛋白表达量分别比前一梯度低浓度干预组有显著升高(P0.05),NGF、LIF上调IL-4、IL-5mRNA和蛋白表达呈浓度依赖性。结论:在哮喘大鼠中,NGF、LIF可能通过上调Th2类细胞因子IL-4、IL-5mRNA表达和蛋白分泌,参与促进和放大Th2类细胞因子免疫失衡效应。     

A role for leukemia inhibitory factor in melanoma-induced bone metastasis

Melanoma is commonly associated with multi-organ metastasis, and bone is a frequent metastatic site for melanoma. However, the mechanism responsible for such melanoma-induced bone metastasis is still poorly understood. In the present study, the intracardiac inoculation of leukemia inhibitory factor (LIF)-producing human melanoma-derived cells (SEKI) developed osteolytic bone destruction in male BALB/cA-nu/nu nude mice. To elucidate the role of LIF in melanoma-induced osteolysis, cells were prepared in which the expression of LIF was reduced using a siRNA technique from the parent SEKI cells. Osteoclastogenesis was induced in the co-culture of LIF and/or SEKI cells with osteoblastic stromal cells in vitro, whereas the LIF-reduced SEKI cells did not induce osteoclastogenesis. The intracardiac inoculation of LIF-reduced SEKI cells resulted in a significant reduction in the incidence and number of bone metastasis in comparison to those in the mice inoculated with the parent SEKI cells. The expression of LIF was found in seven of nine human melanoma-derived cell lines, suggesting that LIF expression is a universal event in melanoma. These findings suggest that a potential role for LIF in the melanoma-induced bone metastasis possibly through the stimulation of osteoclastogenesis. LIF might therefore be a potentially effective drug target in the treatment of bone metastasis in melanoma.     

米非司酮流产蜕膜组织中白血病抑制因子表达的研究

目的探讨早孕妇女米非司酮药物流产与非药物人工流产的蜕膜组织中白血病抑制因子（LIF）表达的差异,分析米非司酮的致流产作用与LIF维持妊娠作用之间的关系。方法运用免疫组织化学技术检测23例米非司酮流产妇女蜕膜组织LIF蛋白的表达,并与20例非药物人工流产蜕膜组织进行对照。结果LIF蛋白在所有蜕膜标本中均有表达,其表达强度在米非司酮流产蜕膜组织和非药物人工流产蜕膜组织中无明显差异（P〉0.05）。结论米非司酮对早孕蜕膜组织中LIF的表达可能无明显影响。     

Expression of interleukin-6 receptor, leukemia inhibitory factor receptor and glycoprotein 130 in the murine cerebellum and neuropathological effect of leukemia inhibitory factor on cerebellar Purkinje cells

Expression of glycoprotein 130 and the related receptors, including interleukin-6 receptor and leukemia inhibitory factor receptor, was examined in the murine cerebellum at the protein level. Western blot analysis revealed that interleukin-6 receptor, leukemia inhibitory factor receptor and glycoprotein 130 were expressed in the murine cerebellum. Immunoreactivities for interleukin-6 receptor, leukemia inhibitory factor receptor and glycoprotein 130 were strongly localized on the cell body of Purkinje cells, indicating that both interleukin-6 and leukemia inhibitory factor could act directly on Purkinje cells in murine adult mice. The expressions of interleukin-6 receptor, leukemia inhibitory factor receptor and glycoprotein 130 were observed on the cell membranes of Purkinje cells by immunoelectron microscopy. Immunoreactivity for the interleukin-6 receptor was also detected in the cytoplasm of Purkinje cells. Injection of a murine hemopoietic cell line, FDC-P1 cells, transfected with the complementary DNA encoding the leukemia inhibitory factor led to a reduction in calbindin-positive dendrites of the Purkinje cells.

The present results suggest that the leukemia inhibitory factor affects cerebellar functions through Purkinje cells.     

Opposing effects of tumor necrosis factor alpha and leukemia inhibitory factor in lipopolysaccharide-stimulated myelopoiesis.

Three myelopoietically active, lipopolysaccharide (LPS)-stimulated monokines, interleukin-1 alpha (IL-1 alpha), tumor necrosis factor alpha (TNF-alpha), and leukemia inhibitory factor (LIF), were tested for effect in an in vitro model for LPS-induced inflammatory murine monocytopoiesis. Neither cytokine stimulated clonal proliferation of marrow-derived progenitors; however, both IL-1 alpha and TNF-alpha enhanced macrophage colony-stimulating factor (M-CSF)-dependent colony formation. The additional progenitors stimulated by IL-1 alpha and TNF-alpha to form colonies in response to M-CSF were equivalent to the precommitment, transitional progenitors stimulated by M-CSF and bacterial LPS. In addition, the additional colonies elicited by IL-1 alpha and TNF-alpha were not additive in cultures containing both M-CSF and LPS, indicating these colonies arose from the same LPS-responsive, two-signal-dependent transitional progenitors. Leukemia inhibitory factor did not influence M-CSF-stimulated colony formation; however, LIF effected a dose-dependent inhibition of colony formation by transitional progenitors responding to combinations of M-CSF and LPS, IL-1 alpha, TNF-alpha, or an additional transitional cell costimulant, substance P. Neutralizing anti-murine TNF-alpha antibodies abrogated transitional cell colony formation stimulated by combinations of M-CSF and TNF-alpha, IL-1 alpha, LPS, or substance P but had no effect on colony formation stimulated solely by M-CSF. The results indicate that TNF-alpha may be an important positive stimulus for commitment of progenitors to the mononuclear phagocyte lineage and that TNF-alpha may be the endogenous regulator of the costimulatory effects of LPS, IL-1, and substance P. In addition, the results indicate that LIF may play an opposing negative regulatory role acting to inhibit LPS and TNF-alpha stimulation of the transitional progenitors.     

Effects of leukemia inhibitory factor on lectin-binding patterns in the uterine stromal vessels of mice

Lectin histochemistry was performed on mouse uteri to determine what effects leukemia inhibitory factor (LIF) has on carbohydrate epitope expressions at the time of implantation. Twenty-two biotinylated lectins were used in this study. Following injection of LIF, specific binding to the apical surface of the uterine glandular epithelium (GE) was recognized by six lectins. Particularly, binding of the lectin from Griffonia (Bandeiraea) simplicifolia was specific to the glandular epithelium close to the luminal epithelium. Succinylated wheat germ agglutinin (WGA), which has specificity for oligosaccharides recognized by WGA without sialic acid residues, showed weaker binding to the uterine luminal epithelium (LE) and the stroma than WGA, suggesting that terminal residues of glyco-conjugates on these tissues may be modified by sialic acids. Lectin binding to the glandular and luminal epithelium was not influenced by LIF. However, three lectins including a lectin from Dolichos biflorus showed specificity for stromal vessels 6h after LIF injection. Since the lectin from D. biflorus binds to neo-vascular vessels, LIF may play a role in regulating maternal angiogenesis directly and/or indirectly during implantation.     

白血病抑制因子在妊娠输卵管黏膜中的表达

目的探讨白血病抑制因子(LIF)与输卵管妊娠的关系。方法选择输卵管妊娠组25例、正常输卵管组28例及正常宫内早孕组30例,采用免疫组织化学染色技术及半定量病理图像分析系统检测输卵管妊娠黏膜组织、正常输卵管黏膜组织及正常宫内早孕组子宫蜕膜组织中LIF的表达。结果 LIF阳性表达在正常宫内早孕组最高,在输卵管妊娠组中较高,在正常输卵管组中最低,三组两两比较差异均有有统计学意义(P<0.05)。结论 LIF可能参与了输卵管妊娠发病过程,且与输卵管妊娠缺乏蜕膜化反应有关。     

Localization of the murine leukemia inhibitory factor gene near the centromere on chromosome 11

Leukemia inhibitory factor (LIF) is a glycoprotein with divergent activities: It induces the differentiation of certain myeloid leukemic cells, inhibits the differentiation of embryonic stem cells, and promotes bone remodelling in vivo and in vitro. The murine LIF gene has been assigned to the proximal region of chromosome 11 at sub-bands A1-A2, by analysis of a panel of mouse x Chinese hamster somatic cell hybrids and by in situ hybridization. Interestingly, the proximal portion of chromosome 11 has been shown, by virtue of its parental origin effects, to contain gene(s) involved in fetal growth. It is also interesting that there is a preponderance of chromosome 11 abnormalities in embryonal carcinoma cells. The localization of the murine LIF gene confirms the homology of a portion of murine chromosome 11 with human chromosome 22q, the site of the human LIF gene.     

Detection of human leukemia inhibitory factor by monoclonal antibody based ELISA.

Leukemia inhibitory factor (LIF) is known to exhibit multiple functions by regulating the growth and differentiation of multiple normal cell types as well as malignant cells. To have a better understanding of the role of LIF, it is important to determine the level of LIF in various biological samples by developing an easy, sensitive and LIF specific assay. In this study, we have established a double monoclonal antibody (mAb) based ELISA. Four hybridoma cell lines (D3.14.1, D4.16.9, D25.1.4 and D62.3.2) secreting murine monoclonal antibodies (mAbs) against recombinant human leukemia inhibitory factor (rHuLIF) were produced by immunization of BALB/c mice with rHuLIF and by fusing immune spleen cells with P3X63Ag8U.1 myeloma cells. These mAbs each belong to the IgG1 isotype and have unique isoelectrofocusing point patterns. All four mAbs were shown to have high affinities for rHuLIF (Kd = 7 x 10(-10) to 6 x 10(-11) M) and were able to recognize the native as well as the reduced rHuLIF in an immunoblotting assay. All these mAbs showed no cross-reactivities to IL-1, IL-3, IL-6, TNF-alpha, GCSF and GMCSF. MAb D3.14.1 showed a weak binding to Oncostatin M but not to rMuLIF whereas the other three mAbs D4.16.9, D25.1.4 and D62.3.2 showed cross-reactivity to rMuLIF but not to Oncostatin M. Data obtained from a competitive binding enzyme-linked immunosorbent assay (ELISA) suggested that these four mAbs recognized different epitopes on rHuLIF. Using mAb D4.16.9 as coat antibody and horseradish peroxidase (HRP) conjugated mAb D3.14.1 as the conjugate antibody we established a double mAb based ELISA specific for human LIF which could detect as little as 100 pg/ml and 10 pg/ml of rHuLIF in the absence and in the presence of the ELAST ELISA amplification system, respectively. The addition of serum had very minimal effect on this ELISA.     

Gestational vitamin B deficiency leads to homocysteine-associated brain apoptosis and alters neurobehavioral development in rats

Hyperhomocysteinemia has been identified as a risk factor for neurological disorders. To study the influence of early deficiency in nutritional determinants of hyperhomocysteinemia on the developing rat brain, dams were fed a standard diet or a diet lacking methyl groups during gestation and lactation. Homocysteinemia progressively increased in the offspring of the deficient group and at 21 days reached 13.3+/-3.7 micromol/L versus 6.8+/-0.3 micromol/L in controls. Homocysteine accumulated in both neurons and astrocytes of selective brain structures including the hippocampus, the cerebellum, the striatum, and the neurogenic subventricular zone. Most homocysteine-positive cells expressed p53 and displayed fragmented DNA indicative of apoptosis. Righting reflex and negative geotaxis revealed a delay in the onset of integration capacities in the deficient group. Between 19 and 21 days, a poorer success score was recorded in deficient animals in a locomotor coordination test. A switch to normal food after weaning allowed restoration of normal homocysteinemia. Nevertheless, at 80 days of age, the exploratory behavior in the elevated-plus maze and the learning and memory behavior in the eight-arm maze revealed that early vitamin B deprivation is associated with persistent functional disabilities, possibly resulting from the ensuing neurotoxic effects of homocysteine.     

Granulocyte factor and graft-versus-host reaction in rats. I. The inhibitory effect of granulocyte factor on the local graft-versus-host reaction in rats

Recent data have proved that PMNLs regulate the immune responses in vitro and in vivo. PMNL specific granules contain an immunoregulatory factor termed the granulocyte factor (GF). The glycogen-induced PMNL infiltration to peritoneal cavity of rats significantly diminished the local GvHR. GF, injected subcutaneously three times at a day of challenge, one and two days after, significantly diminished local GvHR as well. GF treatment before parental lymphocyte injection had no effect on the local GvHR in rats. Inactivation of GF using anti-GF IgG antibodies, abolished the inhibitory effect of PMNLs in GvHR.     

Influence of age on iminodipropionitrile-induced vestibular and neurobehavioral toxicities in rats

A direct association between aging and drug-induced dyskinesia has been reported by several investigators. Iminiodipropionitrile (IDPN), a prototype nitrile compound produces a motor syndrome in rodents, which resembles neuroleptic drug induced dyskinesia. In this investigation attempt has been made to study the effect of age on IDPN induced vestibular hair cell degeneration and resulting dyskinetic syndrome. Male Wistar rats aged 3, 6 and 12 weeks received IDPN in the doses of 0, 200 and 400 mg/kg, intraperitoneally for 3 consecutive days. IDPN-induced dyskinesia was assessed using a behavioral testing battery on days 3, 4, 5, 6, 7, 14, 21 and 28. The rats were sacrificed on day 28; temporal bones were excised for vestibular histopathology and sera were collected for measuring the indices of oxidative stress (glutathione and conjugated dienes). IDPN in the dose of 200 mg/kg produced dyskinesia in 12 weeks old rats, but failed to do so in 3 and 6 weeks old rats. The high dose of IDPN (400 mg/kg) caused dyskinesia in all age groups, however, its onset and severity were age-dependent. Older rats showed an early onset and significantly high incidence of dyskinesia as compared to younger rats. The susceptibility of rats to IDPN-induced behavioral deficits was proportional to oxidative stress and degeneration of sensory hair cells in the crista ampullaris.     

Macrophage migration inhibitory factor expression in male and female ethanol-fed rats.

Macrophage migration inhibitory factory (MIF) regulates macrophage accumulation at sites of injury and can promote the inflammatory response. We studied MIF expression in the intragastric feeding rat model for alcoholic liver injury. Male and age-matched female rats were fed ethanol or dextrose with fish oil. Two groups of male rats were fed medium-chain triglycerides with ethanol or dextrose. Analysis of liver histopathology, lipid peroxidation, endotoxin, mRNA, and immunohistochemistry for MIF, tumor necrosis factor-alpha (TNF-alpha) and interferon-gamma (IFN-gamma) were carried out. Male and female rats fed fish oil and ethanol showed necroinflammatory liver injury and had the highest expression of MIF, TNF-alpha, and IFN-gamma in the liver. Decreased levels of MIF protein were seen in rats with higher endotoxin levels, suggesting that preformed MIF is released into the circulation. MIF is an important mediator of the inflammatory response in alcoholic liver disease and a potential therapeutic target.     

Effect of leukemia inhibitory factor on hemopoietic and stromal precursor cells in prolonged culture of mouse bone marrow

Treatment of prolonged bone marrow cultures with leukemia inhibitory factor during the first 2 weeks after explantation has no appreciable effect on the production of precursors and mature hemopoietic cells during 4 weeks of culturing. However, the proliferative potential of polypotent hemopoietic precursors in these cultures increases substantially. The addition of exogenous cytokine has a pronounced effect on the hemopoietic stroma, specifically, on the content of osteogenic precursors and cells transporting the hemopoietic microenvironment to prolonged bone marrow cultures treated by leukemia inhibitory factor. This effect is confirmed by formation of ectopic hemopoietic fociin vivo, being 2–3 times higher than in the control. Translated fromByulleten' Eksperimental'noi Biologii i Meditsiny, Vol. 122, No. 9, pp. 325–328, September, 1996