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.

Analysis of Intravenous Insulin Dosing Requirements for Treatment of Severe Hypertriglyceridemia.

Purpose: Insulin infusion therapy is commonly utilized for treatment of severe hypertriglyceridemia, however, data supporting a standardized dosing approach is lacking. This study aimed to determine the average initial insulin dose utilized for treatment of hypertriglyceridemia and the associated reduction in serum triglycerides. Methods: This single-center, retrospective, descriptive analysis conducted at an academic medical center included adult hospitalized patients with serum triglyceride levels greater than 1000 mg/dL receiving treatment with an intravenous insulin infusion between November 2017 and August 2020. Data was extracted from the electronic medical record. The primary outcome was the mean weight-based intravenous insulin dose resulting in resolution of hypertriglyceridemia. Secondary outcomes included time to resolution of hypertriglyceridemia, adverse events associated with insulin treatment, incidence of rebound hypertriglyceridemia, and use of additional lipid-lowering therapies. Results: Data from 32 hospital encounters was analyzed. The mean initial triglyceride level was 3229 mg/dL. The average insulin doses observed on days 1 and 2 of therapy were 0.07 and 0.05 units/kg/hour, respectively. The mean percent triglyceride reduction at 48 hours was 40%. Mean time to resolution of hypertriglyceridemia, discontinuation of insulin infusion, or discharge was 5.7 days. Hypoglycemia and hypokalemia were observed in 9% and 29% of patients, respectively. Conclusion: The results of this study provide new guidance for insulin dosing for hypertriglyceridemia. Serum potassium levels and blood glucose should be monitored closely during therapy.

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.

Vulval lichen sclerosus: An Australasian management consensus.

BACKGROUND/OBJECTIVES: Vulval lichen sclerosus (VLS) is a chronic inflammatory skin condition predominantly affecting the anogenital region in women and children. To date, there is lack of agreement amongst experts on a severity scale to aid assessment, research and treatment stratification on VLS. Furthermore, literature on best practice for long-term management of VLS is lacking. The aim of this consensus is to provide broad guidelines on the short and long-term management of VLS. METHODS: An initial focus group of Australasian experts in vulval dermatology developed a draft consensus statement for the management of VLS. Based on the results of the draft statement, a consensus panel of 22 Australasian experts, comprised of the initial and additional members, participated in an anonymous four-stage eDelphi process. Round 1 involved generation and voting on statements from the draft consensus statement developed by the focus group. In Rounds 2, 3 & 4, panel members were presented formal feedback from previous rounds and asked to indicate their level of agreement. Consensus was reached if there was ≥70% agreement on the importance of an item in the 4 (agree) to 5 (strongly agree) range. RESULTS: The expert panel, with a total of 504 collective years of experience in the field of VLS, reached consensus on a core set of 51 management statements related to diagnosis, severity, initial and long-term management, follow-up, and complications of VLS. CONCLUSIONS: This study has identified a set of management statements for VLS that may be useful in clinical practice in the Australasian population.

Interactions between vulvovaginal disorders and urinary disorders: The case for an integrated view of the pelvis.

Lower urinary tract symptomatology is often difficult to categorize if history and investigation focus only on the urinary tract. Disease and dysfunction in organs more posteriorly can often cause or influence such bladder and urethral symptoms. Vulvovaginal skin diseases are an important but often missed influence on lower urinary tract symptomatology.

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.

The aetiology of chronic vulval pain and entry dyspareunia: a retrospective review of 525 cases.

BACKGROUND: There are few published data about the incidence of diagnoses or treatment outcomes, for chronic vulval pain. AIMS: To document diagnoses and treatment outcomes in a cohort of chronic vulval pain presentations. MATERIALS AND METHODS: A retrospective case review of the patient database of a private vulval clinic between January 2011 and March 2015. RESULTS: Five hundred and twenty-five out of 3360 patients (15.6%) met the criterion of vulval pain alone. Mean age was 47.1 years (range 17-86). Average duration of symptoms was 60 months (range 3-432). Overall, 277/525 (52.7%) patients had satisfactory responses to appropriate treatment and 90/525 (17%) had partial improvement. A dermatosis was identified in 322/525 (61.3%) patients and of these, 211/322 (65.5%) had satisfactory responses to appropriate dermatological treatment. In the remaining 203/525 (38.7%) the skin was normal. These patients were questioned around the possibility of a neuromuscular cause for their pain, including pre-existing dysfunction, trauma or previous operations involving the spine, hips or lower limbs. There were 181/203 (89%) patients considered to have a neuromuscular cause for their pain and considered suitable for physiotherapy and/or neuromodulating medications. Of these patients, 63/182 (34.6%) had satisfactory responses to this treatment. One hundred and sixty-six out of 525 (31.6%) described vulval pain only during sexual intercourse. There was no statistically significant difference between different diagnoses and responses to treatment between patients reporting dyspareunia only and those sexually active women who did not experience dysparenunia (29/525, 5.5%). CONCLUSIONS: The majority of this cohort with chronic vulval pain had a dermatological disease with a smaller proportion caused by neuromuscular dysfunction. Both groups are potentially treatable.

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.

Vulvovaginal candidiasis as a chronic disease: diagnostic criteria and definition.

OBJECTIVE: Although recurrent vulvovaginal candidiasis is defined as 4 or more discrete attacks of vulvovaginal candidiasis per year, there is no diagnostic nomenclature or definition for the many women who are chronically symptomatic. This study aims to establish and propose a definition and a set of diagnostic criteria, which would enable clinicians to promptly identify and treat women with chronic vulvovaginal candidiasis (CVVC). DESIGN: Prospective cohort study. SETTING: Public and private vulvar dermatology outpatient clinics in Sydney, Australia. PARTICIPANTS: Data were obtained prospectively from 50 women with presumptive CVVC and 42 controls. Historical and clinical features of CVVC identified by expert consensus were compared between the 2 groups. Diagnostic criteria were then prospectively applied to a further 163 patients to verify their accuracy. OUTCOME MEASURES: Signs and symptoms diagnostic of CVVC. RESULTS: The following characteristics were found to be significantly more common in women with CVVC compared to controls (p ≤ .001): a history of positive vaginal Candida swab, discharge, dyspareunia, soreness, swelling, cyclicity, and exacerbation of symptoms with antibiotics. CONCLUSIONS: We propose that CVVC can be confidently diagnosed using the major criteria of a chronic nonspecific and nonerosive vulvovaginitis that includes at least 5 or more properties from the following criteria: soreness, dyspareunia, positive vaginal swab either at presentation or in the past, previous response to antifungal medication, exacerbation with antibiotics, cyclicity, swelling, and discharge. This condition responds reliably to oral antifungal medication.