Isolating salient variations of interest in single-cell data with contrastiveVI.

Single-cell datasets are routinely collected to investigate changes in cellular state between control cells and the corresponding cells in a treatment condition, such as exposure to a drug or infection by a pathogen. To better understand heterogeneity in treatment response, it is desirable to deconvolve variations enriched in treated cells from those shared with controls. However, standard computational models of single-cell data are not designed to explicitly separate these variations. Here, we introduce contrastive variational inference (contrastiveVI; https://github.com/suinleelab/contrastiveVI ), a framework for deconvolving variations in treatment-control single-cell RNA sequencing (scRNA-seq) datasets into shared and treatment-specific latent variables. Using three treatment-control scRNA-seq datasets, we apply contrastiveVI to perform a variety of analysis tasks, including visualization, clustering and differential expression testing. We find that contrastiveVI consistently achieves results that agree with known ground truths and often highlights subtle phenomena that may be difficult to ascertain with standard workflows. We conclude by generalizing contrastiveVI to accommodate joint transcriptome and surface protein measurements.

Effect of assisted reproductive technology on multiple sclerosis relapses: Case series and meta-analysis.

BACKGROUND: Five case series reported increased relapse risk after assisted reproductive technologies (ART) in women with multiple sclerosis (MS), but small numbers and heterogeneous study design limit broader conclusions. OBJECTIVE: To evaluate the risk of relapses after ART in an independent case series and in aggregated analyses of existing studies. METHODS: We compared annualized relapse rate (ARR) in the 3 months after, and 12 months before, ART in (1) an unpublished cohort (Boston: prospectively collected relapses; 22 ART cycles), (2i) data pooled from Boston and five published studies (164 cycles), and (2ii) a meta-analysis of all case series published by 2017 (220 cycles; PRISMA and MOOSE guidelines). RESULTS: In the Boston cohort, mean ARR was not higher after ART than before (mean: 0.18 ± 0.85 vs 0.27 ± 0.55, p = 0.58). In the pooled analyses, ARR was significantly higher after ART for all clinical scenarios, including varying ART protocols (p ⩽ 0.01 for each). The meta-analysis confirmed an increased ARR after ART (mean difference (MD) = 0.92, 95% confidence interval (CI) = [0.33, 1.51], p = 0.01). CONCLUSION: These pooled data support an increase in ARR following ART. Reasons for local variation in ARR after ART, and consideration of MS treatments during conception attempts, will be pursued.

Nucleoid and cytoplasmic localization of small RNAs in Escherichia coli.

Bacterial small RNAs (sRNAs) regulate protein production by binding to mRNAs and altering their translation and degradation. sRNAs are smaller than most mRNAs but larger than many proteins. Therefore it is uncertain whether sRNAs can enter the nucleoid to target nascent mRNAs. Here, we investigate the intracellular localization of sRNAs transcribed from plasmids in Escherichia coli using RNA fluorescent in-situ hybridization. We found that sRNAs (GlmZ, OxyS, RyhB and SgrS) have equal preference for the nucleoid and cytoplasm, and no preferential localization at the cell membrane. We show using the gfp mRNA (encoding green fluorescent protein) that non-sRNAs can be engineered to have different proportions of nucleoid and cytoplasmic localization by altering their length and/or translation. The same localization as sRNAs was achieved by decreasing gfp mRNA length and translation, which suggests that sRNAs and other RNAs may enter the densely packed DNA of the nucleoid if they are sufficiently small. We also found that the Hfq protein, which binds sRNAs, minimally affects sRNA localization. Important implications of our findings for engineering synthetic circuits are: (i) sRNAs can potentially bind nascent mRNAs in the nucleoid, and (ii) localization patterns and distribution volumes of sRNAs can differ from some larger RNAs.

Horizontal transfer of DNA methylation patterns into bacterial chromosomes.

Horizontal gene transfer (HGT) is the non-inherited acquisition of novel DNA sequences. HGT is common and important in bacteria because it enables the rapid generation of new phenotypes such as antibiotic resistance. Here we show that in vivo and in vitro DNA methylation patterns can be horizontally transferred into bacterial chromosomes to program cell phenotypes. The experiments were performed using a synthetic system in Escherichia coli where different DNA methylation patterns within the cis-regulatory sequence of the agn43 gene turn on or off a fluorescent reporter (CFP). With this system we demonstrated that DNA methylation patterns not only accompany the horizontal transfer of genes into the bacterial cytoplasm but can be transferred into chromosomes by: (i) bacteriophage P1 transduction; and (ii) transformation of extracellular synthetic DNA. We also modified the experimental system by replacing CFP with the SgrS small RNA, which regulates glucose and methyl α-D-glucoside uptake, and showed that horizontally acquired DNA methylation patterns can increase or decrease cell fitness. That is, horizontally acquired DNA methylation patterns can result in the selection for and against cells that have HGT. Findings from these proof-of-concept experiments have applications in synthetic biology and potentially broad implications for bacterial adaptation and evolution.

Cutaneous melanoma metastatic to the orbit: review of 15 cases.

PURPOSE: To describe the clinical features, management, and outcomes of 15 patients with cutaneous melanoma metastatic to the orbit. The authors review emerging treatments for metastatic melanoma and their ocular implications. METHODS: Retrospective chart review of 15 patients with orbital metastasis from cutaneous melanoma. RESULTS: At presentation of the orbital metastasis, systemic metastatic cutaneous melanoma was present in 13 (87%) patients. The mean interval from diagnosis of cutaneous melanoma to orbital metastasis was 40 months (median, 37 months; range, 0-117 months). The most common presenting signs were dysmotility (63%), proptosis (56%), and blepharoptosis (19%). Four patients (25%) presented with pain. Metastasis involved extraocular muscle in 6 orbits (35%), intraconal space in 4 (24%), extraconal space in 7 (41%), and lacrimal sac in 1 (6%). The tumor was unifocal in all cases, unilateral in 13 patients (87%), and bilateral in 2 (13%). The mean tumor basal dimension was 20 × 20 mm and mean thickness was 16 mm. Treatments included complete surgical excision in 1 patient (6%), external beam radiotherapy (EBRT) in 7 (47%), systemic chemotherapy in 8 (53%), and immunotherapy in 5 (33%). Orbital tumor control was achieved in 2 orbits (18%) following focal therapy alone (excision or EBRT), 4 (36%) following systemic therapy alone (chemotherapy or immunotherapy), and 3 (27%) following combination focal plus systemic therapy. Three patients required exenteration. Survival rates at 1 year/2 years were 100%/0% following focal therapy, 50%/25% following systemic therapy, and 100%/66% following combination therapy. CONCLUSIONS: Cutaneous melanoma metastatic to the orbit tends to involve muscle (35%) or intraconal soft tissue (24%) as a painless (75%), circumscribed (87%) mass. Treatment with systemic chemotherapy and/or immunotherapy resulted in orbital tumor control in 80% of cases. Overall survival was 25.1 months.

Massive macronodular juvenile xanthogranuloma of the eyelid in a newborn.

A newborn baby with a lump on his right upper eyelid that was unresponsive to warm compresses and oral antibiotics presented at 3 weeks of age with a yellow mass measuring 20 mm in diameter at the base. Preliminary diagnosis was benign choristomatous mass; and warm compresses were continued. The mass continued to enlarge and 5 weeks later was 35 mm, with a tight, atrophic overlying epidermis and greater pupillary occlusion. Concern for possible malignancy prompted surgical resection and reconstruction with a supraclavicular graft. Histopathology disclosed that the eyelid tissue was nearly completely replaced by a highly cellular histiocytic neoplasm with prominent eosinophilic, often foamy cytoplasm, and nuclear pleomorphism. Poorly formed Touton giant cells were found. The mass showed positive immunoreactivity, with histiocytic markers CD163 and factor XIII, and was negative for cytokeratin markers, smooth muscle actin, and desmin. These features were compatible with JXG.

Calcified scleral choristoma in organoid nevus syndrome simulating retinoblastoma.

A child referred for management of retinoblastoma who alternatively had a calcified scleral choristoma as part of previously undiagnosed organoid nevus syndrome is described. A 31-month-old male infant with scalp alopecia was referred for retinoblastoma management after a calcified mass in his left eye was found. Ophthalmic examination revealed the mass was of choroidal or scleral origin, underlying the retina. The amelanotic circumpapillary mass extended superonasally in a geographic configuration and measured 14×12 mm. There was no subretinal fluid, hemorrhage, feeder vessels, or tumor seeding. Ocular ultrasonography confirmed a homogeneous calcified intraocular mass 3.1 mm in thickness. Enhanced depth imaging optical coherence tomography revealed that the lesion was located within the sclera compressing the overlying choroidal tissue. Further evaluation disclosed cutaneous aplasia cutis congenita with nevus sebaceous of Jadassohn. Magnetic resonance imaging disclosed an arachnoid cyst of the brain. Later, optical coherence tomography revealed the mass to be in the deep choroid or within the sclera. This constellation of ocular, cutaneous, and neurological features were suggestive of organoid nevus syndrome. At the 2-year follow-up, the findings were stable. The calcified choristoma of organoid nevus syndrome, located within the sclera in this case, has distinctive clinical features that differentiate this benign tumor from retinoblastoma.

Fabrication and mechanical evaluation of anatomically-inspired quasilaminate hydrogel structures with layer-specific formulations.

A major tissue engineering challenge is the creation of multilaminate scaffolds with layer-specific mechanical properties representative of native tissues, such as heart valve leaflets, blood vessels, and cartilage. For this purpose, poly(ethylene glycol) diacrylate (PEGDA) hydrogels are attractive materials due to their tunable mechanical and biological properties. This study explored the fabrication of trilayer hydrogel quasilaminates. A novel sandwich method was devised to create quasilaminates with layers of varying stiffnesses. The trilayer structure was comprised of two "stiff" outer layers and one "soft" inner layer. Tensile testing of bilayer quasilaminates demonstrated that these scaffolds do not fail at the interface. Flexural testing showed that the bending modulus of acellular quasilaminates fell between the bending moduli of the "stiff" and "soft" hydrogel layers. The bending modulus and swelling of trilayer scaffolds with the same formulations were not significantly different than single layer gels of the same formulation. The encapsulation of cells and the addition of phenol red within the hydrogel layers decreased bending modulus of the trilayer scaffolds. The data presented demonstrates that this fabrication method can make quasilaminates with robust interfaces, integrating layers of different mechanical properties and biofunctionalization, and thus forming the foundation for a multilaminate scaffold that more accurately represents native tissue.