Production of a p65fl/fl/LysMCre mouse model with dysfunctional NF-κB signaling in bone marrow-derived macrophages.

Here, we describe the production and characterization of a novel p65fl/fl/LysMCre mouse model, which lacks canonical nuclear factor-kappaB member RelA/p65 (indicated as p65 hereafter) in bone marrow-derived macrophages. Cultured bone marrow-derived macrophages that lack p65 protein reveal NF-κB signaling deficiencies, a reduction in phagocytic ability, and reduced ability to produce nitrites. Despite abnormal bone marrow-derived macrophage function, p65fl/fl/LysMCre mice do not exhibit differences in naïve systemic immune profiles or colony forming units and time to death following Salmonella infection as compared to controls. Additionally, p65fl/fl/LysMCre mice, especially females, display splenomegaly, but no other obvious physical or behavioral differences as compared to control animals. As bone marrow-derived macrophages from this transgenic model are almost completely devoid of canonical nuclear factor-kappaB pathway member p65, this model has the potential for being very useful in investigating bone marrow-derived macrophage NF-kappaB signaling in diverse biological and biomedical studies.

Development of a novel purification protocol to isolate and identify brain microglia.

Microglia, the tissue-resident macrophage of the central nervous system (CNS), play a paramount role in brain health and disease status. Here, we describe a novel method for enriching and isolating primary microglia from mouse brain tissue. This isolation method yields a high number of cells from either young or adult mice, and importantly, maintains the health and function of the cells for subsequent cell culture. We also describe flow cytometry methods using novel cell surface markers, including CX3CR1 and Siglec-H, to specifically label microglia while avoiding other bone marrow and/or non-CNS derived macrophages and monocytes, which has been historically difficult to achieve. As microglia are crucial in multiple aspects of biology, such as in normal brain development/function, immune response, neurodegeneration, and cancer, this isolation technique could greatly benefit a wide range of studies in human CNS biology, health, and disease mechanisms. Being able to isolate a largely pure population of microglia could also allow for a more comprehensive understanding of their functional dynamics and role in disease mechanisms, advancement of potential biomarkers, and development of novel therapeutic targets to improve prognosis and quality of life in multiple diseases.

Lichen sclerosus in pregnancy: A review of 33 cases.

Vulval lichen sclerosus (VLS) is a chronic inflammatory skin condition affecting the anogenital area in women. Serious long-term consequences of VLS include the risk of developing squamous cell carcinoma of the vulva as well as of scarring and alteration of vulval architecture. The treatment of choice for genital lichen sclerosus in females is potent to very potent topical corticosteroids. There are few published data on the course of VLS in pregnancy. We present our experience of managing 33 pregnancies in 29 women with VLS.

Cyproterone acetate in the treatment of oestrogen hypersensitivity vulvovaginitis.

Oestrogen hypersensitivity vulvovaginitis is a rare chronic cyclical vulvovaginitis responsive to oestrogen suppression or antagonism. We present a case series of 16 patients with refractory cyclical vulvovaginitis, all of whom responded to oestrogen suppression with cyproterone acetate.

Canonical NFκB signaling in myeloid cells is required for the glioblastoma growth.

Tumor development and therapeutic resistance are linked with tumor-associated macrophage (TAM) and myeloid-derived suppressor cell (MDSC) infiltration in tumors via chemokine axis. Chemokine expression, which determines the pro or anti-inflammatory status of myeloid cells, are partly regulated by the nuclear factor-kappa B (NF-κB) pathway. Here, we identified that conditional deletion of canonical NF-κB signaling (p65) in myeloid cells inhibited syngeneic glioblastoma (GBM) through decreased CD45 infiltration in tumors, as characterized by decreased TAMs (CD206+) and MDSCs (Gr1+ CD11b+), increased dendritic cells (CD86+) and cytotoxic T cells (CD8+) in the p65 knockout (KO) mice. Proinflammatory cytokines (IFNγ, MCP1, MIP1α, and TNFα) and myeloid differentiation factor (Endoglin) were increased in myeloid cells from p65 KO tumor, which demonstrated an influence on CD8+T cell proliferation. In contrast, p65KO athymic chimeric mice with human GBM, failed to inhibit tumor growth, confirming the contribution of T cells in an immune competent model. The analysis of human datasets and GBM tumors revealed higher expression of p65 in GBM-associated CD68+ macrophages compared to neighboring stroma. Thus, canonical NF-κB signaling has an anti-inflammatory role and is required for macrophage polarization, immune suppression, and GBM growth. Combining an NF-κB inhibitor with standard therapy could improve antitumor immunity in GBM.

Anti-Jagged Immunotherapy Inhibits MDSCs and Overcomes Tumor-Induced Tolerance.

Myeloid-derived suppressor cells (MDSC) are a major obstacle to promising forms of cancer immunotherapy, but tools to broadly limit their immunoregulatory effects remain lacking. In this study, we assessed the therapeutic effect of the humanized anti-Jagged1/2-blocking antibody CTX014 on MDSC-mediated T-cell suppression in tumor-bearing mice. CTX014 decreased tumor growth, affected the accumulation and tolerogenic activity of MDSCs in tumors, and inhibited the expression of immunosuppressive factors arginase I and iNOS. Consequently, anti-Jagged therapy overcame tumor-induced T-cell tolerance, increased the infiltration of reactive CD8+ T cells into tumors, and enhanced the efficacy of T-cell-based immunotherapy. Depletion of MDSC-like cells restored tumor growth in mice treated with anti-Jagged, whereas coinjection of MDSC-like cells from anti-Jagged-treated mice with cancer cells delayed tumor growth. Jagged1/2 was induced in MDSCs by tumor-derived factors via NFkB-p65 signaling, and conditional deletion of NFkB-p65 blocked MDSC function. Collectively, our results offer a preclinical proof of concept for the use of anti-Jagged1/2 to reprogram MDSC-mediated T-cell suppression in tumors, with implications to broadly improve the efficacy of cancer therapy. Cancer Res; 77(20); 5628-38. ©2017 AACR.

Ovulatory disorders are an independent risk factor for pregnancy complications in women receiving assisted reproduction treatments.

BACKGROUND: Conception using assisted reproduction treatments (ART) has been associated with an increased risk of pregnancy complications. It is uncertain if this is caused by ART directly, or is an association of the underlying factors causing infertility. AIMS: We assessed the relationship between assisted conception (AC) and maternal or fetal complications in a large retrospective cohort study. In a nested cohort of women receiving infertility treatment, we determined if such risk rests predominantly with certain causes of infertility. MATERIALS AND METHODS: Retrospective database analysis of 50 381 women delivering a singleton pregnancy in four public hospital obstetric units in western Sydney, and a nested cohort of 508 women receiving ART at a single fertility centre, in whom the cause of infertility was known. RESULTS: A total of 1727 pregnancies followed AC; 48 654 were spontaneous conceptions. Adjusted for age, body mass index and smoking, AC was associated with increased risk of preterm delivery (OR 1.73, 95% CI 1.50-2.02), hypertension (OR 1.55, 95% CI 1.34-1.82) and diabetes (OR 1.51, 95% CI 1.30-1.75). In the nested cohort, ovulatory dysfunction was present in 145 women and 336 had infertility despite normal ovulatory function. Ovulatory dysfunction was associated with increased risk of diabetes (OR 2.94, 95% CI 1.72-5.02) and hypertension (OR 2.40, 95% CI 1.15-5.00) compared to women with normal ovulatory function. CONCLUSIONS: Assisted conception is associated with increased risk of pregnancy complications. This risk appears greatest for women whose underlying infertility involves ovulatory dysfunction. Such disorders probably predispose towards diabetes and hypertension, which is then exacerbated by pregnancy.

A survey of fertility and sexual health following allogeneic haematopoietic stem cell transplantation in New South Wales, Australia.

Four hundred and twenty-one adult allogeneic haematopoietic stem cell transplant (HSCT) survivors participated in a cross-sectional study to assess sexual dysfunction and infertility post-transplant. Survey instruments included the Sydney Post-Blood and Marrow Transplant (BMT) Survey, Functional Assessment of Cancer Treatment (FACT) - BMT, the Depression, Anxiety, Stress Scales (DASS 21), the Chronic Graft-versus-Host Disease (cGVHD) Activity Assessment- Patient Self Report (Form B), the Lee cGVHD Symptom Scale and The Post-Traumatic Growth Inventory. Most HSCT survivors reported sexual difficulties (51% of males; 66% of females). Men reported erectile dysfunction (79%) and decreased libido (61·6%) and women reported loss of libido (83%), painful intercourse (73%) and less enjoyment of sex (68%). Women also commonly reported vaginal dryness (73%), vaginal narrowing (34%) and vaginal irritation (26%). Woman had much higher rates of genital cGvHD than men (22% vs. 5%). Age and cGVHD were significantly associated with sexual dysfunction. Few survivors had children following transplant (3·3%). However, for those of reproductive age at HSCT, 22% reported trying to conceive, with 10·3% reporting success. This study is the largest to date exploring sexual function in survivors of allo-HSCT. This data provides the basis for health service reform to better meet the needs of HSCT survivors, including evidence to support counselling and education both pre- and post-transplant.

Long-term Management of Adult Vulvar Lichen Sclerosus: A Prospective Cohort Study of 507 Women.

IMPORTANCE: Adult vulvar lichen sclerosis (VLS) may be complicated by loss of vulvar structure and vulvar carcinoma. There is a lack of evidence as to the ideal method to maintain long-term remission and prevent complications. OBJECTIVES: To determine whether long-term preventive topical corticosteroid (TCS) treatment of VLS, with a target outcome of induction and maintenance of normal skin texture and color, reduces the risk of vulvar carcinoma, relieves symptoms, improves function, and preserves vulvar architecture, and to evaluate the adverse effects of treatment. DESIGN, SETTING, AND PARTICIPANTS: A prospective longitudinal cohort study was conducted in 507 women with biopsy-proved VLS from January 2, 2008, through September 26, 2014, in the private practice of a dermatologist and a gynecologist in Sydney, Australia. INTERVENTIONS: Preventive treatment using TCSs of various potencies, adjusted to meet a target outcome of normal skin color and texture, with regular long-term follow-up by a dermatologist or gynecologist. MAIN OUTCOMES AND MEASURES: Symptoms or signs of VLS, scarring, development of malignant neoplasms, and adverse effects. RESULTS: The mean age at presentation was 55.4 years (range, 18-86 years); duration of symptoms at presentation, 5.0 years (range, 0.1-40.0 years); and duration of follow-up, 4.7 years (range, 2.0-6.8 years). Remission was induced with a potent TCS, followed by regular preventive TCS treatment of a potency titrated to achieve the target outcome. Patients were followed up at least annually. A total of 150 patients (29.6%) did not carry out the advised treatment and were considered partially compliant. A total of 357 patients (70.4%) adhered to treatment instructions and were considered compliant. Biopsy-proved squamous cell carcinoma or vulvar intraepithelial neoplasia occurred during follow-up in 0 of the compliant patients vs. 7 (4.7%) of the partially compliant patients (P < .001). Suppression of symptoms occurred in 333 (93.3%) compliant patients vs. 87 (58.0%) partially compliant patients (P < .001). Adhesions and scarring occurred during follow-up in 12 (3.4%) compliant patients and 60 (40.0%) partially compliant patients (P < .001). Reversible TCS-induced cutaneous atrophy occurred in 4 (1.1%) compliant patients and 3 (2.0%) partially compliant patients. CONCLUSIONS AND RELEVANCE: This prospective, single-center, longitudinal cohort study of adult patients with VLS suggests that individualized preventive TCS regimens that achieve objective normality of skin color and texture and are used by compliant patients who attend regular long-term follow-up visits may modify the course of the disease. There was a significant difference in symptom control, scarring, and occurrence of vulvar carcinoma between compliant and partially compliant patients. The adverse effects of TCSs were minimal.

IKK/nuclear factor-kappaB and oncogenesis: roles in tumor-initiating cells and in the tumor microenvironment.

The IKK/nuclear factor-kappaB pathway (NF-κB) is critical in proper immune function, cell survival, apoptosis, cellular proliferation, synaptic plasticity, and even memory. While NF-κB is crucial for both innate and adaptive immunity, defective regulation of this master transcriptional regulator is seen in a variety of diseases including autoimmune disease, neurodegenerative disease, and, important to this review, cancer. While NF-κB functions in cancer to promote a number of critical oncogenic functions, here we discuss the importance of the NF-κB signaling pathway in contributing to cancer through promotion of the tumor microenvironment and through maintenance/expansion of tumor-initiating cells, processes that appear to be functionally interrelated.

Surgical division of labial adhesions in vulvar lichen sclerosus and lichen planus.

OBJECTIVE: Vulvar lichen sclerosus (LS) and lichen planus (LP) may cause persistent symptomatic labial adhesions. In the scant literature on this topic, there is no agreement about which operation is suitable, or the role of suppressive medical therapy. We report on simple perineotomy in the context of careful preoperative and postoperative medical suppressions. MATERIALS AND METHODS: Thirty-five patients were identified within a referral vulvar practice, with symptomatic labial adhesions due to LS or LP. After sharp dissection of adhesions and injection of anesthesia, patients doubled the frequency of their preoperative therapy and underwent close surveillance until complete healing had occurred. Suppression of the inflammatory process was continued indefinitely with regular review. RESULTS: Mean age was 57 years. Of the patients, 27 had LS and 8 had LP. Of the 35 patients, 28 (80%) had dyspareunia or apareunia. Mean symptom duration was 9 years. Of the 35 patients, 21 had posterior fusion, 11 had anterior fusion, and 3 had both anterior and posterior fusions. Of the 35 patients, 17 had mild fusion, 11 had moderate fusion, and 7 had severe introital stenosis. At the 3-month review, 31 of the 35 patients had no refusion. Mean duration of follow-up was 2 years (range = 3 months to 7.5 years). Of the 35 patients, 29 had no late refusion during this time. Of the 18 patients with dyspareunia, 8 had no pain, and 9 had less pain. Of the 10 patients with apareunia, 1 could have sex without pain, and 6 could have sex but with pain. CONCLUSION: Simple perineotomy is adequate to treat persistent labial adhesions, provided that the inflammatory process is carefully suppressed.

Management of vulvovaginal lichen planus: a new approach.

OBJECTIVE: This study aimed to report on a novel approach to therapy in a large private dermatogynecology practice using multimodal therapies with adjunctive use of systemic agents where necessary. MATERIALS AND METHODS: This was a retrospective audit of the presentation and management of 131 patients with a clinical diagnosis of vulvovaginal lichen planus. RESULTS: The most frequently presenting symptoms were genital soreness, itch, and burning. Of the 131 patients, 39 (30%) had extragenital disease, mainly oral. Eighty-four (64%) had no external disease. Twenty-two (17%) had introital erosions as the only visible abnormality. Fifty-five (42%) had some degree of labial fusion. Two had full-thickness vulval intraepithelial neoplasia (VIN). Remission induction was achieved in most patients with superpotent topical steroids, but 53 (40%) of 131 patients used oral prednisolone either as an adjunct therapy or alone. All compliant patients achieved symptomatic and objective disease control in a mean of 7.5 weeks. Of the 131 patients, 48 (37%) required multimodal therapy to maintain their initial improvement. Forty-five (34.3%) patients used topical tacrolimus, usually with topical corticosteroids, for maintenance. Eleven (8.5%) required low-dose weekly methotrexate. Fourteen patients experienced adverse reactions severe enough to lead to the cessation of that treatment. The mean length of follow-up was 6.4 years (range = 1 mo to 15 y). The 2 patients with VIN at presentation have had no recurrence. No other patient has yet developed VIN or carcinoma. CONCLUSIONS: Long-term symptomatic and objective control of vulvovaginal lichen planus is possible but requires multimodal therapies, flexible treatment programs, and the judicious use of oral agents.

An outbreak of necrotizing enterocolitis associated with norovirus genotype GII.3.

BACKGROUND: Necrotizing enterocolitis (NEC) is an acute abdominal emergency of unknown etiology predominantly affecting preterm infants. We describe a cluster of NEC in a level III NICU involving 15 infants over a6-month period. Cohorting and stringent infection control measures were associated with termination of the cluster. A case-control study was used to investigate potential risk factors associated with development of NEC. METHODS: Stool samples were collected from 55 infants (10 of 15 NEC and 45 non-NEC controls). Enteric pathogens were identified by culture and/or molecular diagnostic techniques. For the case-control study, controls were selected from admitted neonates during the same time and in the preceding 6-month period, matched for gestation and birthweight. RESULTS: Forty percent (4/10) of NEC infants had norovirus RNA detected compared with 9% (4/45) of non-NEC infants (OR: 6.83, 95% CI: 1.3-34.9,P = 0.021). A lower rate of prolonged rupture of membranes and a higher rate of maternal smoking was also observed in NEC infants than in controls. No significant differences in incidences of chorioamnionitis, intrapartum antibiotics,volume of feedings, time of first formula feeding, and rates of patent ductus arteriosus or intrauterine growth retardation were detected. CONCLUSIONS: Infants who developed NEC had an increased incidence of norovirus detection in their stool following diagnosis. This further strengthens the case for an etiologic role of norovirus in the pathogenesis of NEC.

Polyglutamine toxicity in non-neuronal cells.

The neurodegenerative polyglutamine diseases are caused by an expansion of unstable polyglutamine repeats in various disease proteins. Although these mutant proteins are expressed ubiquitously in neuronal and non-neuronal cells, they cause selective degeneration of specific neuronal populations. Recently, increasing evidence shows that polyglutamine disease proteins also affect non-neuronal cells. However, it remains unclear how the expression of polyglutamine proteins in non-neuronal cells contributes to the course of the polyglutamine diseases. Here, we discuss recent findings about the expression of mutant polyglutamine proteins in non-neuronal cells and their influence on neurological symptoms. Understanding the contribution of non-neuronal polyglutamine proteins to disease progression will help elucidate disease mechanisms and also help in the development of new treatment options.

Topical immunosuppressants, genital lichen sclerosus and the risk of squamous cell carcinoma: a case report.

BACKGROUND: Squamous cell carcinoma (SCC) has a recognized association with lichen sclerosus (LS) of the vulva. Several recent reports have indicated the usefulness of the new macrolide immunosuppressant agents pimecrolimus and tacrolimus in the treatment of LS, emphasizing the advantage over topical corticosteroids of lack of atophogenicity. Despite this there may be risks involved that could outweigh this benefit. The potential of these medications to potentiate the risk of SCC in LS in the short and long-term is unknown. Once lichen sclerosus is well controlled, there is often no need for ongoing use of superpotent corticosteroids, and there may be no reason to use immunosuppressants when moderate-strength corticosteroids provide adequate control. CASE: A 73-year-old woman with a 10-year history of hypertrophic LS and genital psoriasis presented with intractable superimposed inflammatory vulvitis. She was treated with topical pimecrolimus 1% cream on the assumption that she was either allergic to or intolerant of topical corticosteroids. One month after commencing therapy, she suddenly developed a rapidly growing vulvar tumor. This was excised and proved to be a well-differentiated squamous cell carcinoma. CONCLUSION: It may be safest to restrict the use of topical immunosuppressives to patients with LS who are unable to use topical corticosteroids because of the risk of potentiating SCC.