Contraceptive counseling for iPLEDGE and its burden on dermatologists.

Since 2006, iPLEDGE, a risk evaluation and mitigation strategy (REMS), has attempted to prevent fetal exposures in people taking isotretinoin through contraceptive requirements and regular pregnancy testing. There has been criticism of iPLEDGE's requirements, results, and accessibility. iPLEDGE has placed significant burdens on physicians, patients, and administrative staff. Some level of burden is acceptable to prevent fetal exposures, but iPLEDGE burdens are so strenuous that physicians may choose not to prescribe isotretinoin because of them. There are several evidence-based adaptations that iPLEDGE and physicians can enact to improve the isotretinoin experience. First, physicians can practice shared-decision making in contraceptive counseling and educate patients on long-acting reversible contraceptives (LARCs) to improve the counseling process and outcomes. Second, physicians can take advantage of the reimbursed iPLEDGE contraceptive counseling sessions and refer patients accordingly. Finally, iPLEDGE should recognize the variation in efficacy among contraceptives. Specifically, LARCs and permanent surgical sterilization should be exempt from certain iPLEDGE requirements such as monthly pregnancy testing and attestations. iPLEDGE should work with dermatologists for the continual improvement of iPLEDGE. Communication, repetitive reassessment, and subsequent adaptations will result in better care for patients requiring isotretinoin.

Biochemical-free enrichment or depletion of RNA classes in real-time during direct RNA sequencing with RISER.

The heterogeneous composition of cellular transcriptomes poses a major challenge for detecting weakly expressed RNA classes, as they can be obscured by abundant RNAs. Although biochemical protocols can enrich or deplete specified RNAs, they are time-consuming, expensive and can compromise RNA integrity. Here we introduce RISER, a biochemical-free technology for the real-time enrichment or depletion of RNA classes. RISER performs selective rejection of molecules during direct RNA sequencing by identifying RNA classes directly from nanopore signals with deep learning and communicating with the sequencing hardware in real time. By targeting the dominant messenger and mitochondrial RNA classes for depletion, RISER reduces their respective read counts by more than 85%, resulting in an increase in sequencing depth of 47% on average for long non-coding RNAs. We also apply RISER for the depletion of globin mRNA in whole blood, achieving a decrease in globin reads by more than 90% as well as an increase in non-globin reads by 16% on average. Furthermore, using a GPU or a CPU, RISER is faster than GPU-accelerated basecalling and mapping. RISER's modular and retrainable software and intuitive command-line interface allow easy adaptation to other RNA classes. RISER is available at https://github.com/comprna/riser .

Profiling age and body fluid DNA methylation markers using nanopore adaptive sampling.

DNA methylation plays essential roles in regulating physiological processes, from tissue and organ development to gene expression and aging processes and has emerged as a widely used biomarker for the identification of body fluids and age prediction. Currently, methylation markers are targeted independently at specific CpG sites as part of a multiplexed assay rather than through a unified assay. Methylation detection is also dependent on divergent methodologies, ranging from enzyme digestion and affinity enrichment to bisulfite treatment, alongside various technologies for high-throughput profiling, including microarray and sequencing. In this pilot study, we test the simultaneous identification of age-associated and body fluid-specific methylation markers using a single technology, nanopore adaptive sampling. This innovative approach enables the profiling of multiple CpG marker sites across entire gene regions from a single sample without the need for specialized DNA preparation or additional biochemical treatments. Our study demonstrates that adaptive sampling achieves sufficient coverage in regions of interest to accurately determine the methylation status, shows a robust consistency with whole-genome bisulfite sequencing data, and corroborates known CpG markers of age and body fluids. Our work also resulted in the identification of new sites strongly correlated with age, suggesting new possible age methylation markers. This study lays the groundwork for the systematic development of nanopore-based methodologies in both age prediction and body fluid identification, highlighting the feasibility and potential of nanopore adaptive sampling while acknowledging the need for further validation and expansion in future research.

DECTIN-1: A modifier protein in CTLA-4 haploinsufficiency.

Autosomal dominant loss-of-function (LoF) variants in cytotoxic T-lymphocyte associated protein 4 (CTLA4) cause immune dysregulation with autoimmunity, immunodeficiency and lymphoproliferation (IDAIL). Incomplete penetrance and variable expressivity are characteristic of IDAIL caused by CTLA-4 haploinsufficiency (CTLA-4h), pointing to a role for genetic modifiers. Here, we describe an IDAIL proband carrying a maternally inherited pathogenic CTLA4 variant and a paternally inherited rare LoF missense variant in CLEC7A, which encodes for the ß-glucan pattern recognition receptor DECTIN-1. The CLEC7A variant led to a loss of DECTIN-1 dimerization and surface expression. Notably, DECTIN-1 stimulation promoted human and mouse regulatory T cell (Treg) differentiation from naïve αß and γδ T cells, even in the absence of transforming growth factor-ß. Consistent with DECTIN-1's Treg-boosting ability, partial DECTIN-1 deficiency exacerbated the Treg defect conferred by CTL4-4h. DECTIN-1/CLEC7A emerges as a modifier gene in CTLA-4h, increasing expressivity of CTLA4 variants and acting in functional epistasis with CTLA-4 to maintain immune homeostasis and tolerance.

Recrudescence of Severe Carbamazepine- Induced DRESS Syndrome after Initiation of Levetiracetam.

Drug reaction with eosinophilia and systemic symptoms (DRESS) syndrome is a potentially fatal cutaneous hypersensitivity reaction commonly precipitated by antiepileptic drugs (AEDs). Cross-reactivity among aromatic AEDs is well-documented, but between aromatic and nonaromatic AEDs. We report a patient with severe DRESS syndrome precipitated by aromatic AED carbamazepine with recrudescence approximately 2 weeks after substitution with nonaromatic AED levetiracetam. The patient was treated with high-dose corticosteroids and switched to the benzodiazepine AED clobazam. At follow-up appointment several weeks later, the patient's rash, liver injury, and eosinophilia had resolved.

Safety and efficacy of biological agents in the treatment of Systemic Lupus Erythematosus (SLE).

BACKGROUND: To determine the safety and efficacy of biological agents used in the treatment of systemic lupus erythematosus (SLE) in adults. METHODS: Systematic review and meta-analysis following PRISMA guidelines. DATA SOURCES: MEDLINE (through Pubmed), EMBASE, Cochrane library, Clinicaltrials.gov, Australianclinicaltrials.gov.au, ANZCTR.org.au and WHO International Clinical Trials Registry Platform for studies published from 20 May 2021 and 15 years prior. A grey literature search was performed and completed on 31 May 2021. STUDY CRITERIA: Phase II, III or quasi randomised controlled trials, studies with only cerebral or cutaneous lupus were excluded. DATA EXTRACTION: Two authors independently screened studies for eligibility, extracted, reviewed data for accuracy, and used the Cochrane tool to assess risk of bias. RESULTS: Forty-four studies were identified, consisting of 15 groups of drugs and 25 different biological agents, totalling 16,889 patients. The main outcomes assessed included Systemic Lupus Erythematosus Responder Index (SRI), BILAG-Based Composite Lupus Assessment (BICLA) and combined combined/partial renal remission (CRR/PRR). Four groups of biologics were found to improve outcomes. Anti-interferons: Anifrolumab increased BICLA response and SRI 5 to 8, decreased prednisone dosages, with increased herpes zoster infections, but fewer serious adverse events. Sifalimumab improved SRI but also increased herpes zoster infections. Anti BAFF/BLyS and/or APRIL: Belimumab consistently improved SRI 4, decreased prednisone dosages, increased combined CRR/PRR, and had no adverse safety outcomes. Tabalumab increased SRI 5 at 52 weeks with no steroid sparing effect but was associated with increased infusion related adverse events. Telitacicept improved SRI 4 at 52 weeks, with no increased adverse events, though data was rather sparse. Anti CD-20 monoclonal antibody, Obinutuzumab increased combined CRR/PRR at 1 and 2 years. Anti IL12/23 monoclonal antibody, Ustekinumab, increased SRI 4 to 6, but not BICLA at 24 weeks, with no concerning safety outcomes. CONCLUSION: Multiple biologic agents are shown in high quality studies to have a significant therapeutic impact on outcomes in SLE.

Amyopathic dermatomyositis with TIF1? positivity treated with IVIG.

Dermatomyositis is an inflammatory myopathy involving the skin that typically affects patients between 40-60 years of age and is more likely to be diagnosed in women. Around 10-20% of dermatomyositis cases present with subclinical or absent muscle involvement, termed "clinically amyopathic." Presence of anti-transcription intermediary factor 1? (TIF1?) antibodies is an important indicator of underlying malignancy. We present a patient with anti-TIF1? positive amyopathic dermatomyositis associated with bilateral breast cancer. The patient was safely treated with trastuzumab for breast cancer and intravenous immunoglobulin for dermatomyositis.

Diffuse large B-cell lymphoma and red cell autoimmunity: clinical role and pathogenesis.

Diffuse large B-cell lymphoma (DLBCL) is the most common form of B-cell non-Hodgkin lymphoma (B-NHL) with significant morbidity and mortality despite advancements in treatment. Lymphoma and autoimmune disease both result from breakdowns in normal cell regulatory pathways, and epidemiological studies have confirmed both that B-NHL is more likely to develop in the setting of autoimmune diseases and vice versa. Red cell immunity, as evidenced by direct antiglobulin test (DAT) positivity, has been linked to DLBCL and more recently the pathogenic causes of this association have begun to be better understood using molecular techniques. This project aimed to explore the relationship between red cell autoimmunity and DLBCL. DAT positivity was more common in DLBCL as compared to healthy controls (20.4% vs 3.7%, p=0.0005). Univariate analysis found a non-significant trend towards poorer overall survival in the DAT positive (DAT+) compared to the DAT negative (DAT-) groups (p=0.087). High throughput sequencing was used to compare mutations in DLBCL from DAT+ and DAT- patients. The most frequently mutated genes in 15 patient samples were KMT2D (n=13), MYOM2 (n=9), EP300 (n=8), SPEN (n=7), and ADAMTSL3 (n=7), which were mutated in both DAT+ and DAT- groups. BIRC3 (n=3), FOXO1 (n=3) and CARD11 (n=2) were found to be mutated only in samples from the DAT+ group. These gene mutations may be involved in disease development and progression, and potentially represent targets for future therapy. The immunoglobulin genotype IGHV4-34 is seen more frequently in DLBCL clones than in normal B cells and has intrinsic autoreactivity to self-antigens on red cells, which is largely mediated by two motifs within the first framework region (FR1); Q6W7 and A24V25Y.26 These motifs form a hydrophobic patch which determines red cell antigen binding and are frequently mutated away from self-reactivity in normal B cells. If this does not occur this may provide constant B cell receptor signalling which encourages lymphoma development, a theory known as antigen driven lymphomagenesis. As with previous studies, IGHV4-34 was over-represented (15.6%) in our DLBCL cohort. Furthermore, of 6 IGHV4-34-expressing DLBCL samples five had unmutated hydrophobic patch mutations providing further evidence for antigen-driven lymphomagenesis. Mutation analysis of these five samples demonstrated high frequency of mutations in several genes, including CREBBP and NCOR2. Further research could explore if mutations in CREBBP and NCOR2 work in conjunction with the preserved QW and AVY motifs to promote lymphomagenesis in IGHV4-34-expressing B cells, and if so, could guide future targeted therapy.

Store-and-Forward Images in Teledermatology: Narrative Literature Review.

Background: Store-and-forward (SAF) teledermatology uses electronically stored information, including patient photographs and demographic information, for clinical decision-making asynchronous to the patient encounter. The integration of SAF teledermatology into clinical practice has been increasing in recent years, especially during the COVID-19 pandemic. Despite this growth, data regarding the outcomes of SAF teledermatology are limited. A key distinction among current literature involves comparing the quality and utility of images obtained by patients and trained clinicians, as these metrics may vary by the clinical expertise of the photographer. Objective: This narrative literature review aimed to characterize the outcomes of SAF teledermatology through the lens of patient- versus clinician-initiated photography and highlight important future directions for and challenges of the field. Methods: A literature search of peer-reviewed research was performed between February and April 2021. Key search terms included patient-initiated, patient-submitted, clinician-initiated, clinician-submitted, store-and-forward, asynchronous, remote, image, photograph, and teledermatology. Only studies published after 2001 in English were included. In total, 47 studies were identified from the PubMed electronic database and Google Scholar after omitting duplicate articles. Results: Image quality and diagnostic concordance are generally lower and more variable with patient-submitted images, which may impact their decision-making utility. SAF teledermatology can improve the efficiency of and access to care when photographs are taken by either clinicians or patients. The clinical outcomes of clinician-submitted images are comparable to those of in-person visits in the few studies that have investigated these outcomes. Coinciding with the onset of the COVID-19 pandemic, asynchronous teledermatology helped minimize unnecessary in-person visits in the outpatient setting, as many uncomplicated conditions could be adequately managed remotely via images captured by patients and referring clinicians. For the inpatient setting, SAF teledermatology minimized unnecessary contact during dermatology consultations, although current studies are limited by the heterogeneity of their outcomes. Conclusions: In general, photographs taken by trained clinicians are higher quality and have better and more relevant diagnostic and clinical outcomes. SAF teledermatology helped clinicians avoid unnecessary physical contact with patients in the outpatient and inpatient settings during the COVID-19 pandemic. Asynchronous teledermatology will likely play a greater role in the future as SAF images become integrated into synchronous teledermatology workflows. However, the obstacles summarized in this review should be addressed before its widespread implementation into clinical practice.

Quality and Perceived Usefulness of Patient-Submitted Store-and-Forward Teledermatology Images.

Importance: Patient-submitted images vary considerably in quality and usefulness. Studies that characterize patient-submitted images in a real-life setting are lacking. Objective: To evaluate the quality and perceived usefulness of patient-submitted images as determined by dermatologists and characterize agreement of their responses. Design, Setting, and Participants: This survey study included patient images submitted to the Department of Dermatology at Duke University (Durham, North Carolina) between August 1, 2018, and December 31, 2019. From a total pool of 1200 images, 10 dermatologists evaluated 200 or 400 images each, with every image being evaluated by 3 dermatologists. Data analysis occurred during the year leading up to the article being written. Main Outcomes and Measures: The primary outcomes were the responses to 2 questions and were analyzed using frequency counts and interrater agreement (Fleiss κ) to assess image quality and perceived usefulness. We performed a random-effects logistic regression model to investigate factors associated with evaluators' decision-making comfort. We hypothesized that most images would be of low quality and perceived usefulness, and that interrater agreement would be poor. Results: A total of 259 of 2915 patient-submitted images (8.9%) did not depict a skin condition at all. The final analysis comprised 3600 unique image evaluations. Dermatologist evaluators indicated that 1985 images (55.1%) were useful for medical decision-making and 2239 (62.2%) were of sufficient quality. Interrater agreement for a given image's diagnostic categorization was fair to substantial (κ range, 0.36-0.64), while agreement on image quality (κ range, 0.35-0.47) and perceived usefulness (κ range, 0.29-0.38) were fair to moderate. Senior faculty had higher odds of feeling comfortable with medical decision-making than junior faculty (odds ratio [OR], 3.68; 95% CI, 2.9-4.66; P < .001) and residents (OR, 5.55; 95% CI, 4.38-7.04; P < .001). Images depicting wounds (OR, 1.75; 95% CI, 1.18-2.58; P = .01) compared with inflammatory skin conditions and that were in focus (OR, 5.56; 95% CI, 4.63-6.67; P < .001) had higher odds of being considered useful for decision-making. Conclusions and Relevance: In this survey study including 10 dermatologists, a slight majority of patient-submitted images were judged to be of adequate quality and perceived usefulness. Fair agreement between dermatologists was found regarding image quality and perceived usefulness, suggesting that store-and-forward teledermatology initiatives should consider a physician's individual experiences and comfort level. The study results suggest that images are most likely to be useful when they are in focus and reviewed by experienced attending physicians for wound surveillance, but dermatologists may be burdened by irrelevant or unsuitable images.

A protocol to evaluate immunoglobulin deposits in mouse glomeruli.

Accumulation of immunoglobulin and complement components within the kidneys is a hallmark of glomerulonephritis. Staining and detection of IgG, IgA, IgM, and C3 deposits can assist in diagnosing the underlying causes of nephritis and has implications for the pathological processes underpinning glomerulonephritis. Here, we describe a protocol to detect immune deposits within biological specimens such as mouse kidneys. We detail tissue isolation and processing, immunostaining, and fluorescence microscopy to characterize and quantify the extent of immunological deposits contributing to kidney injury. For complete details on the use and execution of this protocol, please refer to Jiang et al. (2021).

Machine Learning Improves Upon Clinicians' Prediction of End Stage Kidney Disease.

Background and Objectives: Chronic kidney disease progression to ESKD is associated with a marked increase in mortality and morbidity. Its progression is highly variable and difficult to predict. Methods: This is an observational, retrospective, single-centre study. The cohort was patients attending hospital and nephrology clinic at The Canberra Hospital from September 1996 to March 2018. Demographic data, vital signs, kidney function test, proteinuria, and serum glucose were extracted. The model was trained on the featurised time series data with XGBoost. Its performance was compared against six nephrologists and the Kidney Failure Risk Equation (KFRE). Results: A total of 12,371 patients were included, with 2,388 were found to have an adequate density (three eGFR data points in the first 2 years) for subsequent analysis. Patients were divided into 80%/20% ratio for training and testing datasets.ML model had superior performance than nephrologist in predicting ESKD within 2 years with 93.9% accuracy, 60% sensitivity, 97.7% specificity, 75% positive predictive value. The ML model was superior in all performance metrics to the KFRE 4- and 8-variable models.eGFR and glucose were found to be highly contributing to the ESKD prediction performance. Conclusions: The computational predictions had higher accuracy, specificity and positive predictive value, which indicates the potential integration into clinical workflows for decision support.

High-dose, high-frequency ustekinumab therapy for patients with severe hidradenitis suppurativa.

In this case series, ustekinumab therapy demonstrated efficacy in some patients with severe hidradenitis suppurativa previously treated with adalimumab and/or infliximab. Larger prospective studies are needed to evaluate ustekinumab as a treatment option for recalcitrant hidradenitis suppurativa.

P2RY8 variants in lupus patients uncover a role for the receptor in immunological tolerance.

B cell self-tolerance is maintained through multiple checkpoints, including restraints on intracellular signaling and cell trafficking. P2RY8 is a receptor with established roles in germinal center (GC) B cell migration inhibition and growth regulation. Somatic P2RY8 variants are common in GC-derived B cell lymphomas. Here, we identify germline novel or rare P2RY8 missense variants in lupus kindreds or the related antiphospholipid syndrome, including a "de novo" variant in a child with severe nephritis. All variants decreased protein expression, F-actin abundance, and GPCR-RhoA signaling, and those with stronger effects increased AKT and ERK activity and cell migration. Remarkably, P2RY8 was reduced in B cell subsets from some SLE patients lacking P2RY8 gene variants. Low P2RY8 correlated with lupus nephritis and increased age-associated B cells and plasma cells. By contrast, P2RY8 overexpression in cells and mice restrained plasma cell development and reinforced negative selection of DNA-reactive developing B cells. These findings uncover a role of P2RY8 in immunological tolerance and lupus pathogenesis.

Hidradenitis Suppurativa: Host-Microbe and Immune Pathogenesis Underlie Important Future Directions.

Hidradenitis suppurativa (HS) is an inflammatory disease of the skin with a chronic, relapsing-remitting course. The pathogenesis of the disease is poorly understood and involves multiple factors, including genetics, environment, host-microbe interactions, and immune dysregulation. In particular, the composition of the cutaneous microbiome shifts as the disease progresses, although it is unclear whether this is a primary or secondary process. Trials with immunomodulatory therapy elucidate the role of specific immune pathways and cytokine signaling in disease mechanism, such as TNF-α, IL-1ß, IL-12, IL-17, IL-23, and complement. Future studies should continue examining the causes of and contributing factors to microbial changes and immune dysregulation in HS pathogenesis.

Single-Cell RNA Sequencing Reveals Cellular and Transcriptional Changes Associated With M1 Macrophage Polarization in Hidradenitis Suppurativa.

Hidradenitis suppurativa (HS) is a chronic inflammatory skin disease characterized by recurrent abscesses, nodules, and sinus tracts in areas of high hair follicle and sweat gland density. These sinus tracts can present with purulent drainage and scar formation. Dysregulation of multiple immune pathways drives the complexity of HS pathogenesis and may account for the heterogeneity of treatment response in HS patients. Using transcriptomic approaches, including single-cell sequencing and protein analysis, we here characterize the innate inflammatory landscape of HS lesions. We identified a shared upregulation of genes involved in interferon (IFN) and antimicrobial defense signaling through transcriptomic overlap analysis of differentially expressed genes (DEGs) in datasets from HS skin, diabetic foot ulcers (DFUs), and the inflammatory stage of normal healing wounds. Overlap analysis between HS- and DFU-specific DEGs revealed an enrichment of gene signatures associated with monocyte/macrophage functions. Single-cell RNA sequencing further revealed monocytes/macrophages with polarization toward a pro-inflammatory M1-like phenotype and increased effector function, including antiviral immunity, phagocytosis, respiratory burst, and antibody-dependent cellular cytotoxicity. Specifically, we identified the STAT1/IFN-signaling axis and the associated IFN-stimulated genes as central players in monocyte/macrophage dysregulation. Our data indicate that monocytes/macrophages are a potential pivotal player in HS pathogenesis and their pathways may serve as therapeutic targets and biomarkers in HS treatment.