During development, cortical neurons are produced in a temporally regulated sequence from apical progenitors, directly or indirectly, through the production of intermediate basal progenitors. The balance between these major progenitor types is critical for the production of the proper number and types of neurons, and it is thus important to decipher the cellular and molecular cues controlling this equilibrium. Here we address the role of a cell cycle regulator, the CDC25B phosphatase, in this process. We show that, in the developing mouse neocortex of both sex, deleting CDC25B in apical progenitors leads to a transient increase in the production of TBR1+ neurons at the expense of TBR2+ basal progenitors. This phenotype is associated with lengthening of the G2 phase of the cell cycle, the total cell cycle length being unaffected. Using in utero electroporation and cortical slice cultures, we demonstrate that the defect in TBR2+ basal progenitor production requires interaction with CDK1 and is because of the G2 phase lengthening in CDC25B mutants. Together, this study identifies a new role for CDC25B and G2 phase length in direct versus indirect neurogenesis at early stages of cortical development.SIGNIFICANCE STATEMENT This study is the first analysis of the function of CDC25B, a G2/M regulator, in the developing neocortex. We show that removing CDC25B function leads to a transient increase in neuronal differentiation at early stages, occurring simultaneously with a decrease in basal intermediate progenitors (bIPs). Conversely, a CDC25B gain of function promotes production of bIPs, and this is directly related to CDC25B's ability to regulate CDK1 activity. This imbalance of neuron/progenitor production is linked to a G2 phase lengthening in apical progenitors; and using pharmacological treatments on cortical slice cultures, we show that shortening the G2 phase is sufficient to enhance bIP production. Our results reveal the importance of G2 phase length regulation for neural progenitor fate determination.

Although lengthening of the cell cycle and G1 phase is a generic feature of tissue maturation during development, the underlying mechanism remains poorly understood. Here, we develop a time-lapse imaging strategy to measure the four cell cycle phases in single chick neural progenitor cells in their endogenous environment. We show that neural progenitors are widely heterogeneous with respect to cell cycle length. This variability in duration is distributed over all phases of the cell cycle, with the G1 phase contributing the most. Within one cell cycle, each phase duration appears stochastic and independent except for a correlation between S and M phase duration. Lineage analysis indicates that the majority of daughter cells may have a longer G1 phase than mother cells, suggesting that, at each cell cycle, a mechanism lengthens the G1 phase. We identify that the CDC25B phosphatase known to regulate the G2/M transition indirectly increases the duration of the G1 phase, partly through delaying passage through the restriction point. We propose that CDC25B increases the heterogeneity of G1 phase length, revealing a previously undescribed mechanism of G1 lengthening that is associated with tissue development.

P48 Growing burden of dermatophytosis in southern region of Nepal R. Tripathi, V. Paudel, M. Pradhan and B.R. Pandey Grande International Hospital, Kathmandu, Nepal and National Medical College, Birgunj, Nepal Dermatophytosis is becoming challenging to treat, particularly because of its chronicity and recurrence. Studies from India have suggested significant distress being caused to patients socially, emotionally and financially. The aim of the study was to quantify the magnitude of the problem being faced by patients with dermatophytosis in the southern region of Nepal. A cross-sectional, single-time assessment prospective observational study was conducted in the Dermatology and Venereology Department of National Medical College, Birgunj, Nepal (March–April 2019). Of 100 patients, a preponderance of dermatophytosis was found in males (male-to-female ratio 1 5 : 1). Mean patient age was 29 5 years (range 4–75). Mean duration of disease at presentation was 5 months. On average, patients travelled a distance of 28 65 km to hospital. Clinical diagnoses in these patients, in descending order, were tinea corporis (78%), tinea cruris (55%), tinea faciei (22%) and others, with more than one diagnosis in 54%. Body surface area involvement ranged from 1% to 40%. The presence of chronic/recurrent dermatophytosis was noted in one-fourth of patients. Ninety-two per cent of patients were using topical treatment prior to their visit, with the use of a topical overthe-counter steroid in 54%. Ten per cent were using irritant medications such as salicylic acid and 27% had used multiple topicals. Other measures applied by 9% of the studied population included chloroxylenol, ayurvedic ointments, cactus extract, garlic paste and neem oil. Of 87% who gave an estimate of treatment cost, the average was $48. Absence from work (range 1–6 days) was reported by 6%. Moderate-to-severe pruritus affecting sleep was reported by 56%. Complications noted in the study were tinea incognito (15%), lichenification (7%), irritant contact dermatitis, striae, pseudoimbricata and scaling. Positive family history was present in 42%, with 1–6 family members being affected. Fifteen per cent had one or more family members with chronic/recurrent dermatophytosis. The burden of dermatophytosis is high at both personal and societal level in terms of the high cost of treatment, availability and abuse of topical steroid, distress due to pruritus and complications of improper treatment. P49 Toxic epidermal necrolysis in the setting of systemic lupus erythematosus: a case series J. Oldham, A. Ryan, M. Steyn, R. Adams, E. Agius, G. Sanna, M. Fernando, E. Calonje, H. Malhomme de la Roche, E. Benton and D. D’Cruz Guy’s and St Thomas’ NHS Foundation Trust, London, UK There is increased recognition of a characteristic presentation of toxic epidermal necrolysis (TEN) associated with systemic lupus erythematosus (SLE). We present a case series of three patients and discuss aspects of their presentation and outcome. A 35-year-old woman who had SLE since 2017 was admitted with a new-onset rash. Recent medication included prednisolone, esomeprazole and flucloxacillin. She had annular erythematous macules on the trunk, arms and face, evolving to bullous lesions with epidermal detachment of 80% of her body surface area (BSA). She had markers of active lupus and was treated with methylprednisolone, intravenous immunoglobulins (IVIG) and granulocyte colony-stimulating factor. She was discharged and started on belimumab for treatment-resistant SLE. Our second case, a previously well 26year-old woman, was admitted with a skin rash. A toothache was treated with co-codamol, amoxicillin and metronidazole, and she developed a macular erythema to her face, trunk and thighs. This recurred 2 months later and she was treated with penicillin V, flucloxacillin and metronidazole (for Clostridium difficile infection). She was found to have a high antinuclear antibody, double-stranded DNA and low complement, was diagnosed with SLE and treated with steroids, lansoprazole and hydroxychloroquine, and developed a widespread bullous eruption. On transfer to the intensive treatment unit (ITU) she had 70% BSA epidermal detachment. She was treated with intravenous methylprednisolone and IVIG. Her recovery was complicated by recurrent sepsis. Our third case, a 68-year-old woman who had SLE and Sjogren syndrome since 2012, had experienced multiple bullous drug reactions to antibiotics and was on haemodialysis. She presented following a seizure and was treated for pneumonia with clarithromycin. She developed bullae on her buttocks, with multiple dusky patches on her lower limbs. There was no mucosal involvement, with subsequent 20% BSA epidermal detachment. The culprit drugs were thought to be clarithromycin or omeprazole. She was transferred to ITU. Her skin re-epithelialized, but she died of an acute abdomen. It is interesting to note the similarities of presentation in these patients; two had had previous rashes. Proton pump inhibitors and antibiotics were implicated as drug culprits in all. There was no mucosal involvement in any patient. All patients had a skin biopsy, which showed fullthickness epidermal necrosis consistent with TEN, with negative direct immunofluorescence in two patients. In the third patient only, immunofluorescence was consistent with lupus. These patients had a protracted ITU stay of 18, 43 and 16 days, respectively, vs. an average stay of 12 days in our hospital for patients with TEN.

ABSTRACTWhile lengthening of the cell cycle and G1 phase is a generic feature of tissue maturation during development, the underlying mechanism remains still poorly understood. Here we develop a time lapse imaging strategy to measure the four phases of the cell cycle in single neural progenitor cells in their endogenous environment. Our results show that neural progenitors possess a great heterogeneity of the cell cycle length. This duration variability is distributed over all phases of the cell cycle, with the G1 phase being the one contributing primarily to cell cycle variability. Within one cell cycle, each phase duration appears stochastic and independent except for a surprising correlation between S and M phase. Lineage analysis indicates that the majority of daughter cells display longer G1 phase than their mother’s suggesting that at each cell cycle a mechanism lengthens the G1 phase. We identify an actor of the core cell cycle machinery, the CDC25B phosphatase known to regulate G2/M transition, as an indirect regulator of the duration of the G1 phase. We propose that CDC25B acts via a cell to cell increase in G1 phase heterogeneity revealing a novel mechanism of G1 lengthening associated with tissue development.

In the developing neural tube in chicken and mammals, neural stem cells proliferate and differentiate according to a stereotyped spatiotemporal pattern. Several actors have been identified in the control of this process, from tissue-scale morphogens patterning to intrinsic determinants in neural progenitor cells. In a previous study (Bonnet et al. eLife 7, 2018), we have shown that the CDC25B phosphatase promotes the transition from proliferation to differentiation by stimulating neurogenic divisions, suggesting that it acts as a maturating factor for neural progenitors. In this previous study, we set up a mathematical model linking fixed progenitor modes of division to the dynamics of progenitors and differentiated populations. Here, we extend this model over time to propose a complete dynamical picture of this process. We start from the standard paradigm that progenitors are homogeneous and can perform any type of divisions (proliferative division yielding two progenitors, asymmetric neurogenic divisions yielding one progenitor and one neuron, and terminal symmetric divisions yielding two neurons). We calibrate this model using data published by Saade et al. (Cell Reports 4, 2013) about mode of divisions and population dynamics of progenitors/neurons at different developmental stages. Next, we explore the scenarios in which the progenitor population is actually split into two different pools, one of which is composed of cells that have lost the capacity to perform proliferative divisions. The scenario in which asymmetric neurogenic division would induce such a loss of proliferative capacity appears very relevant.

In the developing neural tube in chicken and mammals, neural stem cells proliferate and differentiate according to a stereotyped spatio-temporal pattern. Several actors have been identified in the control of this process, from tissue-scale morphogens patterning (Shh, BMP) to intrinsic determinants in neural progenitor cells. In a previous study (Bonnet et al. eLife 7, 2018), we have shown that the CDC25B phosphatase promotes the transition from proliferation to differentiation in a cell-cycle independent fashion. In this study, we set up a mathematical model linking progenitor modes of division to the dynamics of progenitors and differentiated populations. Here, we build on this previous model to propose a complete dynamical picture of this process. We start from the standard model in which progenitors are homogeneous and can perform any type of divisions (proliferative division yielding two progenitors, asymmetric neurogenic divisions yielding one progenitor and one neuron, and terminal symmetric divisions yielding two neurons). We constraint this model using published data about mode of divisions and population dynamics of progenitors/neurons at different developmental stages (Saade et al. Cell Reports 4, 2013), and check the effect of CDC25B gain of function in this context. Next, we explore the scenarios in which progenitors population is actually split into two different pools, one of which composed of cells that have lost the capacity to perform proliferative divisions (fate restriction). We show that one such scenario appears relevant and calls for further identification of the alternative role of CDC25B in such a fate restriction.

A fundamental issue in developmental biology and in organ homeostasis is understanding the molecular mechanisms governing the balance between stem cell maintenance and differentiation into a specific lineage. Accumulating data suggest that cell cycle dynamics play a major role in the regulation of this balance. Here we show that the G2/M cell cycle regulator CDC25B phosphatase is required in mammals to finely tune neuronal production in the neural tube. We show that in chick neural progenitors, CDC25B activity favors fast nuclei departure from the apical surface in early G1, stimulates neurogenic divisions and promotes neuronal differentiation. We design a mathematical model showing that within a limited period of time, cell cycle length modifications cannot account for changes in the ratio of the mode of division. Using a CDC25B point mutation that cannot interact with CDK, we show that part of CDC25B activity is independent of its action on the cell cycle.

Background Most oligodendrocytes of the spinal cord originate from ventral progenitor cells of the pMN domain, characterized by expression of the transcription factor Olig2. A minority of oligodendrocytes is also recognized to emerge from dorsal progenitors during fetal development. The prevailing view is that generation of ventral oligodendrocytes depends on Sonic hedgehog (Shh) while dorsal oligodendrocytes develop under the influence of Fibroblast Growth Factors (FGFs). Results Using the well-established model of the chicken embryo, we show that ventral spinal progenitor cells activate FGF signaling at the onset of oligodendrocyte precursor cell (OPC) generation. Inhibition of FGF receptors at that time appears sufficient to prevent generation of ventral OPCs, highlighting that, in addition to Shh, FGF signaling is required also for generation of ventral OPCs. We further reveal an unsuspected interplay between Shh and FGF signaling by showing that FGFs serve dual essential functions in ventral OPC specification. FGFs are responsible for timely induction of a secondary Shh signaling center, the lateral floor plate, a crucial step to create the burst of Shh required for OPC specification. At the same time, FGFs prevent down-regulation of Olig2 in pMN progenitor cells as these cells receive higher threshold of the Shh signal. Finally, we bring arguments favoring a key role of newly differentiated neurons acting as providers of the FGF signal required to trigger OPC generation in the ventral spinal cord. Conclusion Altogether our data reveal that the FGF signaling pathway is activated and required for OPC commitment in the ventral spinal cord. More generally, our data may prove important in defining strategies to produce large populations of determined oligodendrocyte precursor cells from undetermined neural progenitors, including stem cells. In the long run, these new data could be useful in attempts to stimulate the oligodendrocyte fate in residing neural stem cells.

During spinal cord development, astrocyte precursors arise from neuroepithelial progenitors, delaminate from the ventricular zone, and migrate toward their final locations where they differentiate. Although the mechanisms underlying their early specification and late differentiation are being deciphered, less is known about the temporal control of their migration. Here, we show that the epithelial-mesenchymal transition regulator Zeb1 is expressed in glial precursors and report that loss of Zeb1 function specifically delays the onset of astrocyte precursor delamination from the ventricular zone, correlating with transient deregulation of the adhesion protein Cadherin-1. Consequently, astrocyte precursor invasion into the Zeb1−/− mutant white matter is delayed, and induction of their differentiation is postponed. These findings illustrate how fine regulation of adhesive properties influences the onset of neural precursor migration and further support the notion that duration of exposure of migrating astrocyte precursors to environmental cues and/or their correct positioning influence the timing of their differentiation.

Journal Article An unusual compressible lower leg plaque Get access E. Agius, E. Agius Department of Dermatology University Hospital Lewisham Lewisham London UK Correspondence: Dr Elaine Agius, Department of Dermatology, University College London Hospitals NHS Foundation Trust, 250 Euston Road, London, NW1 2PQ, UK E‐mail: elaineagius@doctors.net.uk Search for other works by this author on: Oxford Academic Google Scholar J. Ross, J. Ross Department of Dermatology University Hospital Lewisham Lewisham London UK Search for other works by this author on: Oxford Academic Google Scholar E. Calonje, E. Calonje Department of Dermatopathology St John's Institute of Dermatology Guys' and St Thomas's Hospital London UK Search for other works by this author on: Oxford Academic Google Scholar A. Giles, A. Giles Department of Pathology University Hospital Lewisham Lewisham London UK Search for other works by this author on: Oxford Academic Google Scholar S. Morris, S. Morris Department of Oncology Guys' and St Thomas's Hospital London UK Search for other works by this author on: Oxford Academic Google Scholar S. R. Hoque S. R. Hoque Department of Dermatology University Hospital Lewisham Lewisham London UK Search for other works by this author on: Oxford Academic Google Scholar Clinical and Experimental Dermatology, Volume 40, Issue 2, 1 March 2015, Pages 216–218, https://doi.org/10.1111/ced.12482 Published: 01 March 2015

Deciphering the core machinery of the cell cycle and cell division has been primarily the focus of cell biologists, while developmental biologists have identified the signaling pathways and transcriptional programs controlling cell fate choices. As a result, until recently, the interplay between these two fundamental aspects of biology have remained largely unexplored. Increasing data show that the cell cycle and regulators of the core cell cycle machinery are important players in cell fate decisions during neurogenesis. Here, we summarize recent data describing how cell cycle dynamics affect the switch between proliferation and differentiation, with an emphasis on the roles played by the cell cycle regulators, the CDC25 phosphatases.

S BJD British Journal of Dermatology Medical Dermatology Meeting Royal College of Physicians, London Thursday 9 January 2014 Oral presentations Oral 1 Fat necrosis secondary to a deficiency in protein C and antithrombin C. Edwards, J. Uprichard, V. Akhras and E. Agius St George’s Hospital, London, U.K. We present a 19-year-old woman with a background of Down syndrome and Eisenmenger syndrome who presented with a 4-month history of progressive confluent widespread erythema and indurated skin over her abdomen, buttocks and thighs. This was causing pain and considerable difficulty mobilizing. The clinical features were suggestive of a widespread panniculitis. A skin biopsy was taken from the abdomen. This showed the presence of microvascular thrombi and no evidence of panniculitis. However, clinically the findings were still very suggestive of a panniculitis and therefore a second biopsy was performed from the abdominal wall. This again showed no evidence of panniculitis but did show extensive microvascular thrombi and fat necrosis. The fat necrosis was felt to be secondary to thrombotic vascular occlusion within the subcutaneous tissue. A thrombophilia screen showed negative anticardiolipin antibodies and negative lupus anticoagulant, but significantly low protein C and antithrombin levels. These findings were confirmed on three separate sequential blood samples. Our patient was commenced on low-molecular-weight heparin, but finding an adequate site for injection proved problematic, in view of the extensive fat necrosis. She was therefore commenced on oral rivaroxaban as a long-term method of anticoagulation. In conclusion, this is a rare clinical entity whereby a subcutaneous thrombotic vasculopathy resulting in fat necrosis was the only presentation of an underlying protein C and antithrombin deficiency. Oral 2 Acute-onset generalized scleroderma as a feature of chronic graft-versus-host disease (cGVHD): is acute-onset sclerodermatous cGVHD a separate clinical entity? Z. Salih, F. Ferguson and S. Kulkarni The Christie Hospital, Manchester, U.K. Sclerodermatous chronic graft-versus-host disease (cGVHD) is uncommon (10–12%) after allogeneic haematopoietic stem cell transplantation (AHSCT) and classically develops as a slowly progressive event. We describe three cases of scleroderma developing as an acute event. Case 1. One year following AHSCT for Hodgkin lymphoma a patient developed acute scleroderma to the face, trunk and limbs, resulting in restriction of joint mobility. Corticosteroids and mycophenolate were ineffective. Targeted therapy with imatinib and extracorporeal photopheresis (ECP) led to complete resolution of scleroderma. Case 2. One year following AHSCT for Hodgkin lymphoma a patient developed acute scleroderma affecting the trunk and limbs. Corticosteroids, mycophenolate and thalidomide were ineffective. ECP was contraindicated. Methotrexate provided some improvement to significant contractures. Case 3. A patient developed progressive multisystem cGVHD following reduced-intensity AHSCT for multiple myeloma. While on treatment with corticosteroids and mycophenolate, they developed acute scleroderma affecting the trunk, which rapidly became generalized and caused restricted joint mobility and restrictive lung disease. ECP had no benefit. Imatinib was withdrawn as it achieved stabilization only, and thalidomide was discontinued due to sensory neuropathy. Currently, methotrexate has improved sclerodermatous changes. To our knowledge, this is the first report of cGVHD manifesting as acute-onset scleroderma. The clinical features were a combination of eosinophilic fasciitis and classical scleroderma, with no evidence of Raynaud syndrome or autoantibodies. Even though the manifestations are similar, response to therapy and underlying pathogenesis are likely to be variable. Cooperative research efforts between haematology and dermatology are required to improve understanding and to establish whether acute-onset scleroderma should be classified separately. Oral 3 A case of autoimmune lymphoproliferative syndrome E. Orrin, R. Ireland, S. Kiani, I. Mohammed and T.N. Basu Kings College Hospital, London, U.K. A 21-year-old man was referred to dermatology with generalized itch, worsening over 1 year, and intermittent hive-like rashes. Since the age of 3 years, he had been under the care of the paediatric haematologists with unexplained hepatosplenomegaly and intermittent multilineage cytopenias. Aged 21 years the patient had a new diagnosis of autoimmune haemolytic anaemia, which prompted referral to our immunology, haematology and dermatology teams. On examination, the patient had alopecia totalis and vitiligo and was © 2014 British Association of Dermatologists British Journal of Dermatology (2014) 170, ppe7–e13 e7 dermatographic. He had cervical lymphadenopathy and hepatosplenomegaly. The constellation of lymphoproliferation and autoimmune syndromes led our immunologists to suspect autoimmune lymphoproliferative syndrome (ALPS). Immunophenotypic T-cell subset analysis revealed > 5% peripheral double-negative T cells (CD3, CD4 , CD8 , T-cell receptor ab). This cell population is rare in peripheral blood but is found in ALPS. After appropriate counselling, molecular genetic analysis confirmed that he was heterozygous for a C219A mutation of the gene coding for Fas ligand, confirming a diagnosis of ALPS. ALPS is an inherited disorder of abnormal lymphocyte survival caused by defective Fas-mediated apoptosis. It is characterized by the combination of lymphoproliferation (caused by aberrant accumulation of lymphoid cells) and autoimmunity (due to the defective elimination of autoreactive T cells). Prompt diagnosis is important because patients have an increased risk of Hodgkin and non-Hodgkin lymphoma. Hence, management of autoimmune disorders with immunosuppressive therapy on a background of lymphoma susceptibility is a challenge. The patient remains under long-term multidisciplinary follow-up. In conclusion, recognition of the combination of autoimmunity with lymphoproliferation may have hastened the unifying diagnosis of ALPS in our patient, which can otherwise remain elusive.

We investigated the relationship between varicella zoster virus (VZV)-specific memory CD4(+) T cells and CD4(+)Foxp3(+) regulatory T cells (Tregs) that accumulate after intradermal challenge with a VZV skin test Ag. VZV-specific CD4(+) T cells were identified with a MHC class II tetramer or by intracellular staining for either IFN-γ or IL-2 after Ag rechallenge in vitro. VZV-specific T cells, mainly of a central memory (CD45RA(-)CD27(+)) phenotype, accumulate at the site of skin challenge compared with the blood of the same individuals. This resulted in part from local proliferation because >50% of tetramer defined Ag-specific CD4(+) T cells in the skin expressed the cell cycle marker Ki67. CD4(+)Foxp3(+) T cells had the characteristic phenotype of Tregs, namely CD25(hi)CD127(lo)CD39(hi) in both unchallenged and VZV challenged skin and did not secrete IFN-γ or IL-2 after antigenic restimulation. The CD4(+)Foxp3(+) T cells from unchallenged skin had suppressive activity, because their removal led to an increase in cytokine secretion after activation. After VZV Ag injection, Foxp3(+)CD25(hi)CD127(lo)CD39(hi) T cells were also found within the VZV tetramer population. Their suppressive activity could not be directly assessed by CD25 depletion because activated T cells in the skin were also CD25(+). Nevertheless, there was an inverse correlation between decreased VZV skin responses and proportion of CD4(+)Foxp3(+) T cells present, indicating indirectly their inhibitory activity in vivo. These results suggest a linkage between the expansion of Ag-specific CD4(+) T cells and CD4(+) Tregs that may provide controlled responsiveness during Ag-specific stimulation in tissues.