Effects of leukemia inhibitory factor on endogenous neural stem cell proliferation and glycoprotein-130 expression in a mouse model of cerebral infarction

Leukemia inhibitory factor （LIF） has been shown to promote proliferation of endogenous neural stem cells. In this study, we treated mice with cerebral infarction using LIF to investigate whether the LIF receptor subunit glycoprotein （gp）130 is involved in neuroprotection. After LIF treatment, the motor function of model mice was significantly improved. Immunofluorescence histochemistry showed increased numbers of endogenous neural stem cells surrounding the infarct foci. Western blot analysis revealed that gp130 expression was significantly decreased surrounding the infarcted foci. Results demonstrated that LIF promoted the proliferation of endogenous neural stem cells by inhibiting gp130 protein expression.

Effects of leukemia inhibitory factor and basic fibroblast growth factor on free radicals and endogenous stem cell proliferation in a mouse model of cerebral infarction

The present study established a mouse model of cerebral infarction by middle cerebral artery occlusion, and monitored the effect of 25 tJg/kg leukemia inhibitory factor and （or） basic fibroblast growth factor administration 2 hours after model establishment. Results showed that following administration, the number of endogenous neural stem cells in the infarct area significantly increased, malondialdehyde content in brain tissue homogenates significantly decreased, nitric oxide content, glutathione peroxidase and superoxide dismutase activity significantly elevated, and mouse motor function significantly improved as confirmed by the rotarod and bar grab tests. In particular, the effect of leukemia inhibitory factor in combination with basic fibroblast growth factor was the most significant. Results indicate that leukemia inhibitory factor and basic fibroblast growth factor can improve the microenvironment after cerebral infarction by altering free radical levels, improving the quantity of endogenous neural stem cells, and promoting neurological function of mice with cerebral infarction.

Expression of Leukemia Inhibitory Factor in Airway Epithelial Tissue of Asthmatic Rats

In order to investigate the expression of leukemia inhibitory factor （LIF） in airway epithelial tissues of normal and asthmatic rats, the influence of dexamethasone and the role of LIF in pathogenesis of asthma, 30 Sprague-Dawley （SD） rats were randomly divided into 3 groups （10 for each group）： normal group, asthma model group, and dexamethasone-interfered group. In asthma model group and dexamethasone-interfered group, asthma rat models were established by intraperitoneal （i.p.） injection of 10% ovalbumin （OVA） and challenge with 1% OVA via inhalation. Rats in dexamethasone-interfered group were pretreated with dexamethasone （2 mg/kg, i.p） 30 min before each challenge. The expression of LIF protein in lung was detected by immunohistochemistry. The results showed that LIF protein was mainly expressed in cytoplasm of bronchial epithelial cells. The expression of LIF protein in the airway epithelial tissue of asthma model group was significantly higher than that in normal group and dexamethasone-interfered group （P〈0.01）, but there was no significant difference between normal group and dexamethasone-interfered group （P〉0.05）. It was concluded that the expression of LIF was increased significantly in the airway epithelial tissue of the asthma rats, and dexamethasone could down-regulate the expression of LIF. It was suggested that LIF might play an important role in the pathogenesis of asthma as an inflammation regulator.

Leukemia Inhibitory Factor Enhanced the Developmental and Implantation Compatibility of Mouse Embryos in Co-culture with Human Endometrial Epithelial Cells

Objective:Among the variousin vitro embryo culture systems,co-culture has demonstrated remarkable effects in pre-implantation embryo development owing to the production of embryo-nourishing factors.Nevertheless,little is known about the secretion of these factors.Therefore,in this study,the effect of leukemia inhibitory factor(LIF),one of the most important nourishing factors in the early development of mouse embryos,in human endometrial epithelial cells(hEECs)was evaluated.Methods:Two-cell stage embryos were collected from the oviducts of hyper-stimulated and mated mice and cultivated in a co-culture with an hEEC monolayer with or without LIF.The quality and developmental and attachment potential rates of cultured embryos were evaluated by determining the levels of octamer-binding transcription factor 4(Oct4)and caudal type homeobox 2(Cdx2)transcripts.Results:LIF significantly increased the developmental rate(82.67%vs.61.04%,respectively)and attachment rate(64%vs.45.45%,respectively)of mouse embryos co-cultured with hEECs compared to those in untreated embryos.The expression levels ofOct4 andCdx2 in blastocysts cultured in the presence of LIF were higher than those in blastocysts cultured without LIF.Conclusions:Despite the secretion of LIF by hEECs during co-culture with embryos,the amount of this factor was insufficient,and its addition to the culture media could increase the developmental potential of embryos.

Protective Effects of Leukemia Inhibitory Factor on Retinal Vasculature and Cells in Streptozotocin-induced Diabetic Mice

Background： Leukemia inhibitory factor （LIF） has been reported to possess various pharmacological effects, including displaying vascular and neuroprotective properties, during retinal disease. The aim of this study was to investigate the vascular and structural changes in the retina of diabetic mice and to explore whether LIF prevents experimental diabetes-induced retinal injury in the early stages. Methods： Diabetes was induced in C57BI/6J mice with streptozotocin （STZ） injections. Successful diabetic animal models were randomly separated into two groups： the diabetic group （n = 15） and the LIF-treated group （n = 15）. Normal C57BL/6 mice served as the normal control group （n = 14）. Recombinant human LIF was intravitreally injected 8 weeks after the diabetic model was successfully established. Retinas were collected and evaluated using histological and immunohistochemical techniques, and flat-mounted retinas and Western blotting were performed at 18 weeks after the induction of diabetes and 2 days after the intravitreal injection of LIF. The analysis of variance test were used. Results： Histological analysis showed that there were fewer retinal ganglion cells （RGCs） and the inner nuclear layer （INL） became thinner in the diabetic model group （RGC 21.8 ± 4.0 and INL 120.2 ± 4.6 μm） compared with the normal control group （RGC 29.0 ± 6.7, t = -3.02, P = 0.007; INL 150.7 ±10.6 lain, t = -8.88, P 〈 0.001, respectively）. After LIF treatment, the number of RGCs （26.9 ± 5.3） was significantly increased （t = 3.39, P = 0.030） and the INL （ 134.5± 14.2 lain） was thicker compared to the diabetic group （t - 2.75, P = 0.013）. In the anti-Brn-3a-labeled retinas, the number of RGCs in the LIF-treated group （3926.0 ± 143.9） was obviously increased compared to the diabetic group （3507.7 ± 286.1, t = 2.38, P = 0.030）, while no significance was found between the LIF-treated group and the control group （4188.3 ± 114.7, t= -2.47, P- 0.069）. Flat-mounted retinas demonstrated that a disorganized, dense distribution of the vessel was prominent in the diabetic model group. Vessel distribution in the LIF-treated mouse group was typical and the thickness was uniform. The levels of phosphosignal transducer and activator of transcription 3 activation were obviously higher in the LIF-injected retinas than those in the diabetic control group （t = 3.85, P = 0.019） and the normal control （t = -3.20, P - 0.019）. Conclusion： The present study provides evidence that LIF treatment protects the integrity of the vasculature and prevents retinal injury in the early stages of diabetic retinopathy in STZ-induced diabetic models.

Glycemic control predicts the risk of hepatic fibrosis in biopsy-proven NAFLD:a possible mediating role for leukemia inhibitory factor?

Background:Despite the clear link between nonalcoholic fatty liver disease(NAFLD)and type 2 diabetes mellitus,little is understood about how glycemic control impacts histological severity.We aimed to investigate the deeper association between hemoglobin A1c(HbA1c)and the histologic severity of liver fibrosis.Methods:A total of 568 adults with biopsy-proven NAFLD from the PERSONS cohort in Wenzhou were enrolled.The association between mean HbA1c and hepatic histological features was investigated with ordinal logistic regression.Generalized additive models were used to identify the non-linear relationship between HbA1c and increased fibrosis stage(stage F
3).Causal mediation analysis was performed to calculate the indirect effect of the relationship between HbA1c and hepatic fibrosis mediated by cytokines.Results:Every 1%increase in mean HbA1c was associated with 16%higher odds of increased fibrosis stage(odds ratio 1.16,95%CI 1.04–1.30),even after adjustment for confounding factors.There was a non-linear association between HbA1c and increased fibrosis stage(stage F
3),and the HbA1c inflection point was 9.2%.In particular,the ORs on the left and right sides of this inflection point were 2.1(95%CI 1.4–3.3)and 0.1(95%CI 0–4.8),respectively.7%of the association(OR 1.07,95%CI 1.01–1.12)between HbA1c and liver fibrosis was mediated by leukemia inhibitory factor.Conclusions:Glycemic control predicts severity of hepatic fibrosis and leukemia inhibitory factor plays an intermediary role between them,and thus optimizing glycemic control may be a means of modifying the risk of fibrosis progression in NAFLD.

DETECTION OF LEUKEMIA INHIBITORY FACTOR (LIF) BY A ENZYME-LINKED IMMUNOSORBENT ASSAY

LIF is a cytokine with leiotropic activities. In order to understand better the physiological and patho-physiological role of LIF. we have developed a simple and specific enzyme-linked immunosorbent assay (ELISAI for detecting LIF in human plasma and serum and in tissue culture media. A monoclonal ami-LIF antibody 8B11 (IgGl) produced in our laboratory was coated onto microtiter plates. After block-

BMPRⅡ^(+)neural precursor cells isolated and characterized from organotypic neurospheres:an in vitro model of human fetal spinal cord development

Roof plate secretion of bone morphogenetic proteins(BMPs)directs the cellular fate of sensory neurons during spinal cord development,including the formation of the ascending sensory columns,though their biology is not well understood.Type-ⅡBMP receptor(BMPRⅡ),the cognate receptor,is expressed by neural precursor cells during embryogenesis;however,an in vitro method of enriching BMPRⅡ^(+)human neural precursor cells(hNPCs)from the fetal spinal cord is absent.Immunofluorescence was undertaken on intact second-trimester human fetal spinal cord using antibodies to BMPRⅡand leukemia inhibitory factor(LIF).Regions of highest BMPRⅡ^(+)immunofluorescence localized to sensory columns.Parenchymal and meningeal-associated BMPRⅡ^(+)vascular cells were identified in both intact fetal spinal cord and cortex by co-positivity with vascular lineage markers,CD34/CD39.LIF immunostaining identified a population of somas concentrated in dorsal and ventral horn interneurons,mirroring the expression of LIF receptor/CD118.A combination of LIF supplementation and high-density culture maintained culture growth beyond 10 passages,while synergistically increasing the proportion of neurospheres with a stratified,cytoarchitecture.These neurospheres were characterized by BMPRⅡ^(+)/MAP2ab^(+/–)/βⅢ-tubulin^(+)/nestin^(–)/vimentin^(–)/GFAP^(–)/NeuN^(–)surface hNPCs surrounding a heterogeneous core ofβⅢ-tubulin^(+)/nestin^(+)/vimentin^(+)/GFAP^(+)/MAP2ab^(–)/NeuN^(–)multipotent precursors.Dissociated cultures from tripotential neurospheres contained neuronal(βⅢ-tubulin^(+)),astrocytic(GFAP+),and oligodendrocytic(O4+)lineage cells.Fluorescence-activated cell sorting-sorted BMPRⅡ^(+)hNPCs were MAP2ab^(+/–)/βⅢ-tubulin^(+)/GFAP^(–)/O4^(–)in culture.This is the first isolation of BMPRⅡ^(+)hNPCs identified and characterized in human fetal spinal cords.Our data show that LIF combines synergistically with high-density reaggregate cultures to support the organotypic reorganization of neurospheres,characterized by surface BMPRⅡ^(+)hNPCs.Our study has provided a new methodology for an in vitro model capable of amplifying human fetal spinal cord cell numbers for>10 passages.Investigations of the role BMPRⅡplays in spinal cord development have primarily relied upon mouse and rat models,with interpolations to human development being derived through inference.Because of significant species differences between murine biology and human,including anatomical dissimilarities in central nervous system(CNS)structure,the findings made in murine models cannot be presumed to apply to human spinal cord development.For these reasons,our human in vitro model offers a novel tool to better understand neurodevelopmental pathways,including BMP signaling,as well as spinal cord injury research and testing drug therapies.

Synchronous gastric cancer complicated with chronic myeloid leukemia (multiple primary cancers):A case report

BACKGROUND With the advancement of medical technology and improvement in living standards,the incidence of multiple primary cancers has gradually increased.In particular,tumors of the digestive system account for a large proportion of multiple primary cancers.The diagnosis and treatment of chronic myeloid leukemia,particularly with synchronous gastric cancer,at the first consultation is relatively rare.CASE SUMMARY Herein,we present the case of a middle-aged man who was referred to the Department of Hematology owing to an elevated white blood cell count.After the examination,he was diagnosed with chronic myeloid leukemia and was administered imatinib.Three months after the initial diagnosis,he visited our hospital again for abdominal pain,and further examination revealed gastric malignancy.After discussion with a multidisciplinary team,S-1(Tegafur,Gimeracil,and Oteracil Potassium Capsules) combined with oxaliplatin—SOX regimen—was initiated.Later,the patient’s condition rapidly progressed.He developed colonic obstruction and underwent an ostomy;however,he died less than 6 months after the initial diagnosis.CONCLUSION Multiple primary cancers are influenced by environmental and genetic factors;a standardized multidisciplinary discussion plays a key role in treatment.

The neuroprotective role of Wnt signaling in the retina

The canonical Wnt/β-catenin signaling pathway has been shown to play a major role during embryonic development and maturation of the central nervous system including the retina. It has a significant impact on retinal vessel formation and maturation, as well as on the establishment of synaptic structures and neuronal function in the central nervous system. Mutations in components of the Wnt/β-catenin signaling cascade may lead to severe retinal diseases, while dysregulation of Wnt signaling can contribute to disease progression. Apart from the angiogenic role of Wnt/β-catenin signaling, research in the last decades leads to the theory of a protective effect of Wnt/β-catenin signaling on damaged neurons. In this review, we focus on the neuroprotective properties of the Wnt/β-catenin pathway as well as its downstream signaling in the retina.

Protective effect of LIF-huM SCs on the retina of diabetic model rats

AIM: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.METHODS: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(FITCdextran),and retinal structure examination of sections using hematoxylin and eosin staining.Expression levels of adiponectin(APN),high-sensitivity C-reactive protein(hsCRP),and neurotrophin-4(NT-4)in each group was detected using immunohistochemistry,PCR,Western blotting,and ELISA,respectively.RESULTS: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).CONCLUSION:LIF-hUCMSCs protect the retina of diabetic rats by upregulating APN and NT-4 expression and downregulating hs-CRP expression in the retina.

Optical coherence tomography and T cell gene expression analysis in patients with benign multiple sclerosis

Benign multiple sclerosis is a retrospective diagnosis based primarily on a lack of motor symptom progression. Recent findings that suggest patients with benign multiple sclerosis experience non-motor symptoms highlight the need for a more prospective means to diagnose benign multiple sclerosis early in order to help direct patient care. In this study, we present optical coherence tomography and T cell neurotrophin gene analysis findings in a small number of patients with benign multiple sclerosis. Our results demonstrated that retinal nerve fiber layer was mildly thinned, and T cells had a distinct gene expression profile that included upregulation of interleukin 10 and leukemia inhibitory factor, downregulation of interleukin 6 and neurotensin high affinity receptor 1（a novel neurotrophin receptor）. These findings add evidence for further investigation into optical coherence tomography and m RNA profiling in larger cohorts as a potential means to diagnose benign multiple sclerosis in a more prospective manner.

Effects of Jiawei Buzhong Yiqi Decoction on the Endometrial Receptivity of Rats with Polycystic Ovary Syndrome

[Objectives]The study aims to discuss the effects of Jiawei Buzhong Yiqi Decoction on the endometrial receptivity of rats with polycystic ovarian syndrome(PCOS).[Methods]Adult female SD rats were selected and randomly divided into the model group and control group.In the model group,the PCOS model was established by using the method of Poretsky L.Rats in the control group were given physiological saline intervention at the same time.The successfully modeled rats were randomly divided into the PCOS group and traditional Chinese medicine group.Among them,rats in the Chinese medicine group were given Jiawei Buzhong Yiqi Decoction by gavage;rats in the model group and the control group were given normal saline for 14 d.All rats were killed by anesthesia after the last administration,and serum and uterine specimens were collected.Serum sex hormones(including T,E_(2),and P),glucose and insulin levels were detected;the equivalent diameter and area of endometrial gland and gland cavity,and the endometrial thickness of rats were measured;the expression of endometrial leukemia inhibitory factor(LIF),integrinαvβ3 protein and mRNA was detected by using Elisa and RT-PCR methods.[Results]The serum T,glucose,and insulin levels of rats in the PCOS group were significantly higher than those of the control group and the traditional Chinese medicine group,and there was no significant difference in the above indicators between the control group and the traditional Chinese medicine group;there was no significant difference in the E_(2) and P levels between the three groups.The equivalent diameter and area of endometrial gland and gland cavity,and the endometrial thickness of rats in the PCOS group were lower than those of the control group and the traditional Chinese medicine group;there was no significant difference in the above indicators between the traditional Chinese medicine group and the control group.The expression of LIF,integrinαvβ3 protein and mRNA in the endometrium of rats in various groups was basically the same.The expression of LIF and integrinαvβ3 in the endometrium of rats in the PCOS group was significantly lower than that of the control group and the traditional Chinese medicine group;there was no significant difference in the above indicators between the control group and the traditional Chinese medicine group.[Conclusions]The endometrial receptivity of PCOS rats was decreased;Jiawei Buzhong Yiqi Decoction can improve the endometrial receptivity of PCOS rats by increasing the expression of LIF and integrinαvβ3 in the endometrium.

Guizhi Decoction(桂枝汤) Inhibits Cholinergic Transdifferentiation by Regulating Imbalance of NGF and LIF in Salt-Sensitive Hypertensive Heart Failure Rats

Objective: To observe the imbalance of anatomical and functional innervation factors of sympathetic nerves, nerve growth factor(NGF) and leukemia inhibitory factor(LIF), in salt-sensitive hypertensive heart failure rats and to explore the effects of treatment with Guizhi Decoction(桂枝汤) on sympathetic remodeling by inhibiting cholinergic transdifferentiation. Methods: SS-13 BN and Dahl salt-sensitive(DS) rats were divided into 3 groups: SS-13 BN group(control group, n=9), DS group(model group, n=9) and GS group(Guizhi Decoction, n=9). After 10 weeks of a high-salt diet, the GS group rats were given Guizhi Decoction and other two groups were given saline at an equal volume as a vehicle. After 4 weeks’ intragastric administration, rats were executed to detect the relevant indicators. Echocardiography and plasma n-terminal pro-B type natriuretic peptide(NT-proBNP) levels were used to assess cardiac function. Noradrenaline(NA) levels in the plasma and myocardium were detected to evaluate the sympathetic function. NGF and LIF expression were detected in the myocardium by Western blot or quantitative real-time PCR. Double immunofluorescence or Western blot was used to detect tyrosine hydroxylase(TH), choline acetyltransferase(CHAT) and growth associated protein 43(GAP43) in order to reflect anatomical and functional changes of sympathetic nerves. Results: DS group had anatomical and functional deterioration of sympathetic nerves in the decompensation period of heart failure compared with SS-13 BN group. Compared with the DS group, Guizhi Decoction significantly decreased the expression of LIF mRNA/protein(P<0.01), increased the expression of NGF(P<0.05 or P<0.01), enhanced the levels of TH^+/GAP43^+ and TH^+/CHAT^+ positive nerve fibers(P<0.01), and improved the protein expression of TH and GAP43 in left ventricle, but had no effect on CHAT(P>0.05). Guizhi Decoction inhibited inflammatory infiltration and collagen deposition of myocardial injury, increased the content of myocardial NA(P<0.05), reduced the plasma NA level(P<0.01), improved cardiac function(P<0.01), and improved weight and blood pressure to some extent(P<0.05), compared with DS group. Conclusions: Guizhi Decoction could inhibit cholinergic transdifferentiation of sympathetic nerves, improve the anatomical and functional denervation of sympathetic nerves, and delay the progression of decompensated heart failure. The mechanism may be associated with the correction of the imbalance of NGF and LIF.