Single-nucleotide polymorphism array and fluorescence in situ hybridization analysis to decode the cytogenetic profile of atypical partial hydatidiform moles diagnosed by short tandem repeat polymorphism analysis.

Accurate diagnosis of partial hydatidiform moles (PHMs) is crucial for improving outcomes of gestational trophoblastic neoplasia. The use of short tandem repeat (STR) polymorphism analysis to distinguish between PHM and hydropic abortuses is instrumental; however, its diagnostic power has not been comprehensively assessed. Herein, we evaluated the diagnostic efficacy of STR in differentiating between PHM and hydropic abortus, thus providing an opportunity for early measurement of human chorionic gonadotropin for PHMs. We reviewed charts of STR polymorphism analysis performed on fresh villous specimens and patient blood samples using a commercial kit for 16 loci. The genetic classification of 79 PHMs was confirmed. STR was reliable in differentiating PHMs when at least 15 loci were available. Typically, PHMs are characterized by their triploidy, including two paternal and one maternal haploid contribution. In our sample, seven PHMs lacked the three-allelic loci, requiring fluorescence in situ hybridization (FISH) analysis to investigate imbalanced biparental conceptus and single-nucleotide polymorphism array analysis to reveal cytogenetic details. Of these PHMs, two, three, and one were identified as androgenetic/biparental mosaics (diploids), monospermic diandric monogynic triploids, and a typical dispermic diandric monogynic triploid, respectively. The remaining case was monospermic origin, but its ploidy details could not be available. Therefore, STR differentiated PHM from a biparental diploid abortus in most cases. However, PHM diagnosis may be compromised when STR is used as the sole method for cases displaying distinct cytogenetic patterns lacking the three-allelic loci, including androgenetic/biparental mosaicism. Therefore, FISH should be considered to confirm the diagnosis.

STK40 inhibits trophoblast fusion by mediating COP1 ubiquitination to degrade P57Kip2.

BACKGROUND: The syncytiotrophoblast (SCT) layer in the placenta serves as a crucial physical barrier separating maternal-fetal circulation, facilitating essential signal and substance exchange between the mother and fetus. Any abnormalities in its formation or function can result in various maternal syndromes, such as preeclampsia. The transition of proliferative villous cytotrophoblasts (VCT) from the mitotic cell cycle to the G0 phase is a prerequisite for VCT differentiation and their fusion into SCT. The imprinting gene P57Kip2, specifically expressed in intermediate VCT capable of fusion, plays a pivotal role in driving this key event. Moreover, aberrant expression of P57Kip2 has been linked to pathological placental conditions and adverse fetal outcomes. METHODS: Validation of STK40 interaction with P57Kip2 using rigid molecular simulation docking and co-immunoprecipitation. STK40 expression was modulated by lentivirus in BeWo cells, and the effect of STK40 on trophoblast fusion was assessed by real-time quantitative PCR, western blot, immunofluorescence, and cell viability and proliferation assays. Co-immunoprecipitation, transcriptome sequencing, and western blot were used to determine the potential mechanisms by which STK40 regulates P57Kip2. RESULTS: In this study, STK40 has been identified as a novel interacting protein with P57Kip2, and its expression is down-regulated during the fusion process of trophoblast cells. Overexpressing STK40 inhibited cell fusion in BeWo cells while stimulating mitotic cell cycle activity. Further experiments indicated that this effect is attributed to its specific binding to the CDK-binding and the Cyclin-binding domains of P57Kip2, mediating the E3 ubiquitin ligase COP1-mediated ubiquitination and degradation of P57Kip2. Moreover, abnormally high expression of STK40 might significantly contribute to the occurrence of preeclampsia. CONCLUSIONS: This study offers new insights into the role of STK40 in regulating the protein-level homeostasis of P57Kip2 during placental development.

p57Kip2 Phosphorylation Modulates Its Localization, Stability, and Interactions.

p57Kip2 is a member of the cyclin-dependent kinase (CDK) Interacting Protein/Kinase Inhibitory Protein (CIP/Kip) family that also includes p21Cip1/WAF1 and p27Kip1. Different from its siblings, few data are available about the p57Kip2 protein, especially in humans. Structurally, p57Kip2 is an intrinsically unstructured protein, a characteristic that confers functional flexibility with multiple transient interactions influencing the metabolism and roles of the protein. Being an IUP, its localization, stability, and binding to functional partners might be strongly modulated by post-translational modifications, especially phosphorylation. In this work, we investigated by two-dimensional analysis the phosphorylation pattern of p57Kip2 in different cellular models, revealing how the human protein appears to be extensively phosphorylated, compared to p21Cip1/WAF1 and p27Kip1. We further observed clear differences in the phosphoisoforms distributed in the cytosolic and nuclear compartments in asynchronous and synchronized cells. Particularly, the unmodified form is detectable only in the nucleus, while the more acidic forms are present in the cytoplasm. Most importantly, we found that the phosphorylation state of p57Kip2 influences the binding with some p57Kip2 partners, such as CDKs, LIMK1 and CRM1. Thus, it is necessary to completely identify the phosphorylated residues of the protein to fully unravel the roles of this CIP/Kip protein, which are still partially identified.

Chimeric Singleton Placenta Comprising Placental Mesenchymal Dysplasia and Complete Hydatidiform Mole with Live Birth and Postpartum Diagnosis of Gestational Trophoblastic Neoplasia.

INTRODUCTION: Placental mesenchymal dysplasia (PMD) is a benign lesion that is often misdiagnosed as complete (CHM) or partial hydatidiform mole. PMD usually results in live birth but can be associated with several fetal defects. Herein, we report PMD with CHM in a singleton placenta with live birth. CASE PRESENTATION: A 34-year-old gravida 2, para 1, living 1 (G2P1L1) woman was referred on suspicion of a molar pregnancy in the first trimester. Maternal serum human chorionic gonadotrophin levels were increased during early pregnancy, with multicystic lesions and placentomegaly observed on ultrasonography. Levels decreased to normal with no fetal structural abnormalities observed. A healthy male infant was delivered at 34 gestational weeks. Placental p57KIP2 immunostaining and short tandem repeat analysis revealed three distinct histologies and genetic features: normal infant and placenta, PMD, and CHM. Gestational trophoblastic neoplasia was diagnosed and up to fourth-line chemotherapy administered. CONCLUSION: Distinguishing PMD from hydatidiform moles is critical for avoiding unnecessary termination of pregnancy. CHM coexisting with a live fetus rarely occurs. This case is unique in that a healthy male infant was born from a singleton placenta with PMD and CHM.

The significance of CDT1 expression in non-cancerous and cancerous liver in cases with hepatocellular carcinoma.

We previously reported that chromatin licensing and DNA replication factor 1 (CDT1) expression was associated with the extent of proliferation of atypical hepatocytes and the time to postoperative recurrence in cases of hepatocellular carcinoma (HCC). This study aimed to clarify the clinical significance or pathogenesis of CDT1 expression in both non-cancerous and cancerous liver in HCC cases, including previously published data. We investigated the association between the expression of CDT1 in non-cancerous or cancerous liver tissues and histologic findings or biochemical examination results in 62 cases. We also examined the dual localization between CDT1 and FbxW7, P57kip2, P53 and c-Myc by confocal laser scanning microscopy. CDT1 mRNA expression was significantly higher in cancerous liver than in non-cancerous liver (p<0.0001). Elevated CDT1 mRNA expression indicates a significantly degree of inflammatory cell infiltration within lobules, along with elevated serum transaminase levels, and hepatic spare decline. CDT1 mRNA was highly expressed in a group of poorly differentiated cancer cells. CDT1 co-localized with P57kip2, Fbwx7, P53 and c-Myc in the nucleus or cytoplasm of hepatocytes and cancer cells. We found that CDT1 mRNA expression could represent the degree of hepatic spare ability and the high carcinogenic state.

Loss of p57KIP2 expression confers resistance to contact inhibition in human androgenetic trophoblast stem cells.

A complete hydatidiform mole (CHM) is androgenetic in origin and characterized by enhanced trophoblastic proliferation and the absence of fetal tissue. In 15 to 20% of cases, CHMs are followed by malignant gestational trophoblastic neoplasms including choriocarcinoma. Aberrant genomic imprinting may be responsible for trophoblast hypertrophy in CHMs, but the detailed mechanisms are still elusive, partly due to the lack of suitable animal or in vitro models. We recently developed a culture system of human trophoblast stem (TS) cells. In this study, we apply this system to CHMs for a better understanding of their molecular pathology. CHM-derived TS cells, designated as TSmole cells, are morphologically similar to biparental TS (TSbip) cells and express TS-specific markers such as GATA3, KRT7, and TFAP2C. Interestingly, TSmole cells have a growth advantage over TSbip cells only after they reach confluence. We found that p57KIP2, a maternally expressed gene encoding a cyclin-dependent kinase inhibitor, is strongly induced by increased cell density in TSbip cells, but not in TSmole cells. Knockout and overexpression studies suggest that loss of p57KIP2 expression would be the major cause of the reduced sensitivity to contact inhibition in CHMs. Our findings shed light on the molecular mechanism underlying the pathogenesis of CHMs and could have broad implications in tumorigenesis beyond CHMs because silencing of p57KIP2 is frequently observed in a variety of human tumors.

Auxiliary and experimental diagnostic techniques for hydatidiform moles.

Hydatidiform moles are classified into complete hydatidiform moles (CHMs), which are androgenetic and diploid, and partial hydatidiform moles (PHM), which are triploid with two paternal chromosomes and one maternal chromosome. The incidence of gestational trophoblastic neoplasia differs substantially between CHM and PHM. However, they are occasionally difficult to diagnose. In this review, auxiliary and experimental methods based on cytogenetic features and advanced molecular detection techniques applied to the diagnosis and analysis of hydatidiform moles are summarized, including basic principles, characteristics, and clinical implications. Short tandem repeat polymorphism analysis is considered the gold standard for the genetic diagnosis of hydatidiform moles. In clinical settings, immunohistochemical analyses of p57KIP2 , an imprinted gene product, are widely used to differentiate CHMs from other conceptuses, including PHMs. Recently, new molecular genetic techniques, such as single nucleotide polymorphism arrays, have been applied to research on hydatidiform moles. In addition to insights from classical methods, such as chromosome analysis, recently developed approaches have yielded novel findings related to the mechanism underlying the development of androgenetic CHMs.

A Beckwith-Wiedemann-Associated CDKN1C Mutation Allows the Identification of a Novel Nuclear Localization Signal in Human p57Kip2.

p57Kip2 protein is a member of the CIP/Kip family, mainly localized in the nucleus where it exerts its Cyclin/CDKs inhibitory function. In addition, the protein plays key roles in embryogenesis, differentiation, and carcinogenesis depending on its cellular localization and interactors. Mutations of CDKN1C, the gene encoding human p57Kip2, result in the development of different genetic diseases, including Beckwith-Wiedemann, IMAGe and Silver-Russell syndromes. We investigated a specific Beckwith-Wiedemann associated CDKN1C change (c.946 C>T) that results in the substitution of the C-terminal amino acid (arginine 316) with a tryptophan (R316W-p57Kip2). We found a clear redistribution of R316W-p57Kip2, in that while the wild-type p57Kip2 mostly occurs in the nucleus, the mutant form is also distributed in the cytoplasm. Transfection of two expression constructs encoding the p57Kip2 N- and C-terminal domain, respectively, allows the mapping of the nuclear localization signal(s) (NLSs) between residues 220-316. Moreover, by removing the basic RKRLR sequence at the protein C-terminus (from 312 to 316 residue), p57Kip2 was confined in the cytosol, implying that this sequence is absolutely required for nuclear entry. In conclusion, we identified an unreported p57Kip2 NLS and suggest that its absence or mutation might be of relevance in CDKN1C-associated human diseases determining significant changes of p57Kip2 localization/regulatory roles.

Heterogeneous fate choice of genetically modulated adult neural stem cells in gray and white matter of the central nervous system.

Apart from dedicated oligodendroglial progenitor cells, adult neural stem cells (aNSCs) can also give rise to new oligodendrocytes in the adult central nervous system (CNS). This process mainly confers myelinating glial cell replacement in pathological situations and can hence contribute to glial heterogeneity. Our previous studies demonstrated that the p57kip2 gene encodes an intrinsic regulator of glial fate acquisition and we here investigated to what degree its modulation can affect stem cell-dependent oligodendrogenesis in different CNS environments. We therefore transplanted p57kip2 knockdown aNSCs into white and gray matter (WM and GM) regions of the mouse brain, into uninjured spinal cords as well as in the vicinity of spinal cord injuries and evaluated integration and differentiation in vivo. Our experiments revealed that under healthy conditions intrinsic suppression of p57kip2 as well as WM localization promote differentiation toward myelinating oligodendrocytes at the expense of astrocyte generation. Moreover, p57kip2 knockdown conferred a strong benefit on cell survival augmenting net oligodendrocyte generation. In the vicinity of hemisectioned spinal cords, the gene knockdown led to a similar induction of oligodendroglial features; however, newly generated oligodendrocytes appeared to suffer more from the hostile environment. This study contributes to our understanding of mechanisms of adult oligodendrogenesis and glial heterogeneity and further reveals critical factors when considering aNSC mediated cell replacement in injury and disease.

The E2A splice variant E47 regulates the differentiation of projection neurons via p57(KIP2) during cortical development.

During corticogenesis, distinct classes of neurons are born from progenitor cells located in the ventricular and subventricular zones, from where they migrate towards the pial surface to assemble into highly organized layer-specific circuits. However, the precise and coordinated transcriptional network activity defining neuronal identity is still not understood. Here, we show that genetic depletion of the basic helix-loop-helix (bHLH) transcription factor E2A splice variant E47 increased the number of Tbr1-positive deep layer and Satb2-positive upper layer neurons at E14.5, while depletion of the alternatively spliced E12 variant did not affect layer-specific neurogenesis. While ChIP-Seq identified a big overlap for E12- and E47-specific binding sites in embryonic NSCs, including sites at the cyclin-dependent kinase inhibitor (CDKI) Cdkn1c gene locus, RNA-Seq revealed a unique transcriptional regulation by each splice variant. E47 activated the expression of the CDKI Cdkn1c through binding to a distal enhancer. Finally, overexpression of E47 in embryonic NSCs in vitro impaired neurite outgrowth, and overexpression of E47 in vivo by in utero electroporation disturbed proper layer-specific neurogenesis and upregulated p57(KIP2) expression. Overall, this study identifies E2A target genes in embryonic NSCs and demonstrates that E47 regulates neuronal differentiation via p57(KIP2).

Identification of a hydatidiform mole in twin pregnancy following assisted reproduction.

PURPOSE: The aim of this study was to identify a co-existing hydatidiform mole (HM) in twin pregnancy from the abnormal mixed-genomic products of conception (POC) after assisted reproduction by histopathological review, evaluation of p57kip2 immunostaining and short tandem repeat genotyping. METHODS: Thirty-seven patients were collected with suspicion for HM by pathological morphology. They had two embryos individually transferred to their uterus after in vitro fertilization and presented two gestational sacs with undeveloped embryos or one sac with an abnormal area by ultrasonography. RESULTS: Thirty patients were diagnosed as singleton pregnancy, including twenty-two non-molar gestations, six trisomy gestations, one homozygous complete mole and one heterozygous partial mole. Although six patients had ultrasonic imaging of two gestational sacs, the embryonic components in the vacant sac might fade away after transferring. Other seven patients were considered as twin pregnancy by the allelic genotype from two individual conceptions. For the patients with uniform p57kip2 positivity, excessive paternal alleles indicated the potential partial HM in the twin pregnancy. For the patients demonstrated divergent and/or discordant p57kip2 immunostaining, twin pregnancy with co-existing complete HM or mosaic conception were confirmed by genotyping of different villi population respectively. These patients were monitored by serum ß-HCG, while one twin pregnancy with complete mole suffered invasive mole and received chemotherapy. CONCLUSIONS: A strategy composed of selective clinicopathological screening, immunohistochemical interpretation and accurate genotyping is recommended for diagnostically challenging mixed-genomic POC of potential twin pregnancy with HM, especially to differentiate a non-molar mosaic conception from a partial mole.

Postnatal Growth Restriction in Mice Alters Cardiac Protein Composition and Leads to Functional Impairment in Adulthood.

Postnatal growth restriction (PGR) increases the risk for cardiovascular disease (CVD) in adulthood, yet there is minimal mechanistic rationale for the observed pathology. The purpose of this study was to identify proteomic differences in hearts of growth-restricted and unrestricted mice, and propose mechanisms related to impairment in adulthood. Friend leukemia virus B (FVB) mouse dams were fed a control (CON: 20% protein), or low-protein (LP: 8% protein) isocaloric diet 2 weeks before mating. LP dams produce 20% less milk, inducing growth restriction. At birth (postnatal; PN1), pups born to dams fed the CON diet were switched to LP dams (PGR group) or a different CON dam. At PN21, a sub-cohort of CON (n = 3 males; n = 3 females) and PGR (n = 3 males; n = 3 females) were euthanized and their proteome analyzed by two-dimensional differential in-gel electrophoresis (2D DIGE) and mass spectroscopy. Western blotting and silver nitrate staining confirmed 2D DIGE results. Littermates (CON: n = 4 males and n = 4 females; PGR: n = 4 males and n = 4 females) were weaned to the CON diet. At PN77, echocardiography measured cardiac function. At PN80, hearts were removed for western blotting to determine if differences persisted into adulthood. 2D DIGE and western blot confirmation indicated PGR had reductions in p57kip2, Titin (Ttn), and Collagen (Col). At PN77, PGR had impaired cardiac function as measured by echocardiography. At PN80, western blots of p57kip2 showed protein abundance recovered from PN21. PN80 silver staining of large molecular weight proteins (Ttn and Col) was reduced in PGR. PGR reduces cell cycle activity at PN21, which is recovered in adulthood. However, collagen fiber networks are altered into adulthood.

High Dosage Lithium Treatment Induces DNA Damage and p57Kip2 Decrease.

Lithium salt is the first-line therapeutic option for bipolar disorder and has been proposed as a potential antitumoral drug. The effects of LiCl treatment were investigated in SH-SY5Y, a human neuroblastoma cell line and an in vitro model of dopaminergic neuronal differentiation. LiCl, at the dosage used in psychiatric treatment, does not affect cell proliferation, while at higher doses it delays the SH-SY5Y cell division cycle and for prolonged usage reduces cell viability. Moreover, the ion treatment affects DNA integrity as demonstrated by accumulation of p53 and γH2AX (the phosphorylated form of H2AX histone), two important markers of genome damage. p57Kip2, a CIP/Kip protein, is required for proper neuronal maturation and represents a main factor of response to stress including genotoxicity. We evaluated the effect of lithium on p57Kip2 levels. Unexpectedly, we found that lithium downregulates the level of p57Kip2 in a dose-dependent manner, mainly acting at the transcriptional level. A number of different approaches, mostly based on p57Kip2 content handling, confirmed that the CKI/Kip reduction plays a key role in the DNA damage activated by lithium and suggests the unanticipated view that p57Kip2 might be involved in DNA double-strand break responses. In conclusion, our study identified novel roles for p57Kip2 in the molecular mechanism of lithium at high concentration and, more in general, in the process of DNA repair.

P57kip2 immunohistochemical marker as a diagnostic tool for cases of hydatidiform moles in a tertiary health facility in Southwestern Nigeria.

BACKGROUND: Hydatidiform mole (HM) is the most common gestational trophoblastic disease. P57kip2 has been reported to be helpful in differentiating between partial and complete HMs. OBJECTIVES: The study aims to evaluate the P57kip2 immunohistochemical (IHC) marker as a useful ancillary investigation to differentiate complete hydatidiform mole (CHM) from partial hydatidiform mole (PHM). MATERIALS AND METHODS: A retrospective study of all histologically diagnosed HM cases over a 20 year period was undertaken. Clinicopathological parameters were extracted from the surgical day book and medical record archives. Archival haematoxylin- and eosin-stained slides and formalin-fixed paraffin-embedded tissue blocks of all cases of HM diagnosed within the study period were retrieved and reviewed. Cases of HM were reclassified using the P57kip2 IHC marker. The data obtained were analysed using the SPSS version 23. RESULTS: One hundred cases of HMs were studied. CHM accounted for 68%, while PHM accounted for the remaining 32%. The incidence of HM was 2.98 cases per 1000 deliveries. The ratio of CHM to PHM was found to be 2.1:1. Seventy-two per cent of the cases were diagnosed in the first trimester, while the remaining 28% were diagnosed in the second trimester of pregnancy. Based on the P57kip2 IHC staining pattern, HM cases were finally reclassified into 68 cases of CHM and 32 cases of PHM. The age range for all the HM cases was 18-50 years with the majority of the cases seen in the third and fourth decades of life. CONCLUSION: P57kip2 could be useful as an ancillary investigation in confirming the diagnosis of CHM and differentiating it from PHM, particularly in difficult and challenging cases.

Rescuing the negative impact of human endogenous retrovirus envelope protein on oligodendroglial differentiation and myelination.

Remyelination in the adult CNS depends on activation, differentiation, and functional integration of resident oligodendroglial precursor cells (OPCs) and constitutes the only spontaneous neuroregenerative process able to compensate for functional deficits upon loss of oligodendrocytes and myelin sheaths as it is observed in multiple sclerosis. The proteins encoded by p57kip2- and by human endogenous retrovirus type W (pHERV-W) envelope genes were previously identified as negative regulators of OPC maturation. We here focused on the activity of the ENV protein and investigated how it can be neutralized for an improved myelin repair. We could demonstrate that myelination in vitro is severely affected by this protein but that application of an anti-ENV neutralizing antibody, currently investigated in clinical trials, can rescue the generation of internodes. We then compared p57kip2 and ENV dependent inhibitory mechanisms and found that a dominant negative version of the p57kip2 protein can equally save OPCs from myelination failure in response to ENV-mediated TLR4 activation. Additional experiments addressing p57kip2's underlying mode of action revealed a direct interaction with ATP6v1d, a central component of a vascular ATPase. Its pharmacological blocking was then shown to exert an analogous myelination rescue effect in presence of the ENV protein. Therefore, our study provides mechanistic insights into oligodendroglial inhibition processes and presents three different means to counteract the anti-myelination effect of the ENV protein. These observations are therefore of interest in light of understanding the complexity of the numerous oligodendroglial inhibitors and might promote the establishment of novel regenerative therapies.

Long noncoding RNA TALNEC2 plays an oncogenic role in breast cancer by binding to EZH2 to target p57KIP2 and involving in p-p38 MAPK and NF-κB pathways.

We aimed to investigate the potential role and regulatory mechanism of long noncoding RNA tumor-associated lncRNA expressed in chromosome 2 (TALNEC2) in breast cancer. The expression of TALNEC2 in breast cancer tissues and cells were investigated. MCF-7 and MDA-MB-231 cells were transfected with small interfering RNA (siRNA) duplexes for targeting TALNEC2 (si-TALNEC2), enhancer of zeste homolog 2 (EZH2; si-EZH2) and p57KIP2 (si-p57 KIP2 ), and their corresponding controls (si-NC). The viability, colony forming ability, cell cycle, apoptosis, and autophagy of transfected cells were assessed. The expressions of p-p38 mitogen-activated protein kinase (MAPK) and nuclear factor κB (NF-κB) pathway-related proteins were investigated. The results showed that TALNEC2 was highly expressed in breast cancer tissues and cells. Knockdown of TALNEC2 significantly inhibited the malignant behaviors of MCF-7 and MDA-MB-231 cells, including inhibiting cell viability and colony forming, arresting cell cycle at G0/G1 phase, inducing cell apoptosis, and promoting cell autophagy. EZH2 was a TALNEC2 binding protein, which was upregulated in breast cancer tissues and cells and could negatively regulate p57 KIP2 . Effects of TALNEC2 knockdown on malignant behaviors of MCF-7 cells were reversed by p57 KIP2 knockdown. The expressions of p-p38, RelA, and RelB in MCF-7 cells were decreased after knockdown of TALNEC2 or EZH2, which were reversed by knockdown of p57 KIP2 concurrently. In conclusion, TALNEC2 may play an oncogenic role in breast cancer by binding to EZH2 to target p57 KIP2 . Activation of p-p38 MAPK and NF-κB pathways may be key mechanisms mediating the oncogenic role of TALNEC2 in breast cancer. TALNEC2 may serve as a promising target in the therapy of breast cancer.

Teriflunomide promotes oligodendroglial differentiation and myelination.

BACKGROUND: Multiple sclerosis (MS) is a neuroinflammatory autoimmune disease of the central nervous system (CNS) which in most cases initially presents with episodes of transient functional deficits (relapsing-remitting MS; RRMS) and eventually develops into a secondary progressive form (SPMS). Aside from neuroimmunological activities, MS is also characterized by neurodegenerative and regenerative processes. The latter involve the restoration of myelin sheaths-electrically insulating structures which are the primary targets of autoimmune attacks. Spontaneous endogenous remyelination takes place even in the adult CNS and is primarily mediated by activation, recruitment, and differentiation of resident oligodendroglial precursor cells (OPCs). However, the overall efficiency of remyelination is limited and further declines with disease duration and progression. From a therapeutic standpoint, it is therefore key to understand how oligodendroglial maturation can be modulated pharmacologically. Teriflunomide has been approved as a first-line treatment for RRMS in the USA and the European Union. As the active metabolite of leflunomide, an established disease-modifying anti-rheumatic drug, it mainly acts via an inhibition of de novo pyrimidine synthesis exerting a cytostatic effect on proliferating B and T cells. METHODS: We investigated teriflunomide-dependent effects on primary rat oligodendroglial homeostasis, proliferation, and differentiation related to cellular processes important for myelin repair hence CNS regeneration in vitro. To this end, several cellular parameters, including specific oligodendroglial maturation markers, in vitro myelination, and p53 family member signaling, were examined by means of gene/protein expression analyses. The rate of myelination was determined using neuron-oligodendrocyte co-cultures. RESULTS: Low teriflunomide concentrations resulted in cell cycle exit while higher doses led to decreased cell survival. Short-term teriflunomide pulses can efficiently promote oligodendroglial cell differentiation suggesting that young, immature cells could benefit from such stimulation. In vitro myelination can be boosted by means of an early stimulation window with teriflunomide. p73 signaling is functionally involved in promoting OPC differentiation and myelination. CONCLUSION: Our findings indicate a critical window of opportunity during which regenerative oligodendroglial activities including myelination of CNS axons can be stimulated by teriflunomide.

Placental mesenchymal dysplasia with severe fetal growth restriction in one placenta of a dichorionic diamniotic twin pregnancy.

We report a rare case of placental mesenchymal dysplasia (PMD) with fetal growth restriction (FGR) in one placenta of a dichorionic diamniotic (DD) twin pregnancy. A 24-year-old woman was referred to our hospital at 24 weeks' gestation due to FGR and ipsilateral placental abnormality in DD twins. Ultrasound and magnetic resonance imaging showed one placenta of the FGR fetus was bulky and had multiple cysts, while the other fetus placenta appeared normal. Cesarean section was performed at 32 weeks' gestation; the first and second neonates weighted 1799 and 1215 g, respectively. Macroscopically, chorionic vessels on the placental surface of the second neonate were prominently enlarged. Pathological findings demonstrated swelling stem villi with enlarged vessels and increased interstitial cells without trophoblast proliferation. Immunostaining for p57kip2 was negative in interstitial cells and cytotrophoblasts of the swelling stem villi. This suggested that PMD occurred in one placenta of the DD twin, leading to early-onset FGR.

Genetic and Epigenetic Control of CDKN1C Expression: Importance in Cell Commitment and Differentiation, Tissue Homeostasis and Human Diseases.

The CDKN1C gene encodes the p57Kip2 protein which has been identified as the third member of the CIP/Kip family, also including p27Kip1 and p21Cip1. In analogy with these proteins, p57Kip2 is able to bind tightly and inhibit cyclin/cyclin-dependent kinase complexes and, in turn, modulate cell division cycle progression. For a long time, the main function of p57Kip2 has been associated only to correct embryogenesis, since CDKN1C-ablated mice are not vital. Accordingly, it has been demonstrated that CDKN1C alterations cause three human hereditary syndromes, characterized by altered growth rate. Subsequently, the p57Kip2 role in several cell phenotypes has been clearly assessed as well as its down-regulation in human cancers. CDKN1C lies in a genetic locus, 11p15.5, characterized by a remarkable regional imprinting that results in the transcription of only the maternal allele. The control of CDKN1C transcription is also linked to additional mechanisms, including DNA methylation and specific histone methylation/acetylation. Finally, long non-coding RNAs and miRNAs appear to play important roles in controlling p57Kip2 levels. This review mostly represents an appraisal of the available data regarding the control of CDKN1C gene expression. In addition, the structure and function of p57Kip2 protein are briefly described and correlated to human physiology and diseases.

Antagonistic regulation of p57kip2 by Hes/Hey downstream of Notch signaling and muscle regulatory factors regulates skeletal muscle growth arrest.

A central question in development is to define how the equilibrium between cell proliferation and differentiation is temporally and spatially regulated during tissue formation. Here, we address how interactions between cyclin-dependent kinase inhibitors essential for myogenic growth arrest (p21(cip1) and p57(kip2)), the Notch pathway and myogenic regulatory factors (MRFs) orchestrate the proliferation, specification and differentiation of muscle progenitor cells. We first show that cell cycle exit and myogenic differentiation can be uncoupled. In addition, we establish that skeletal muscle progenitor cells require Notch signaling to maintain their cycling status. Using several mouse models combined with ex vivo studies, we demonstrate that Notch signaling is required to repress p21(cip1) and p57(kip2) expression in muscle progenitor cells. Finally, we identify a muscle-specific regulatory element of p57(kip2) directly activated by MRFs in myoblasts but repressed by the Notch targets Hes1/Hey1 in progenitor cells. We propose a molecular mechanism whereby information provided by Hes/Hey downstream of Notch as well as MRF activities are integrated at the level of the p57(kip2) enhancer to regulate the decision between progenitor cell maintenance and muscle differentiation.