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To report the clinicopathologic correlation of a case of bilateral serpiginous-like chorioretinitis (SLC) associated with unilateral ciliochoroidal melanoma.A 71-year-old white woman was diagnosed with progressive SLC in both eyes associated with ciliochoroidal melanoma in the right eye. Clinical findings and imaging before and after enucleation in the right eye were correlated to histologic and immunohistochemistry sections.Examination and imaging identified a peripheral bilobed amelanotic lesion with low reflectivity on B-scan ultrasound with an associated exudative detachment in the right eye. Additionally, multiple areas of new SLC lesions in the macula and peripapillary region in the right eye and along the inferior arcade in the left eye were observed. Oncologic evaluation confirmed a Class 2, ciliochoroidal melanoma, and the eye was enucleated. Autoimmune and infectious laboratory evaluations for the etiology of the SLC lesions were negative. Histopathology of the enucleated eye confirmed the diagnosis of uveal melanoma with lymphocytic inflammation at the edges of the tumor itself and in the areas of discrete SLC lesions. Immunohistochemistry identified similar predominantly CD3 and CD8 T cells and fewer CD20 B cells in both regions.Serpiginous-like chorioretinitis may present as a paraneoplastic, predominantly T-lymphocyte inflammation associated with intraocular tumor such as uveal melanoma.

Adenoid cystic carcinoma of the lacrimal gland is an aggressive, malignant epithelial neoplasm. We report the case of a 30-year-old male with lacrimal gland adenoid cystic carcinoma treated with neoadjuvant intra-arterial chemotherapy through the internal carotid artery, followed by orbital exenteration and chemoradiation. Treatment response was evaluated using a novel combination of pre- and posttreatment genome sequencing coupled with immunohistochemical evaluation, which showed diffuse tumor apoptosis. A posttreatment decrease in variant allele frequency of the NOTCH1 mutation, and robust tumor cytoreduction on imaging, supports exploration of NOTCH1 analysis as a potential marker of cisplatin sensitivity. The use of genome sequencing and immunohistochemical evaluation could provide a more targeted therapeutic assessment of neoadjuvant intra-arterial chemotherapy in the management of lacrimal gland adenoid cystic carcinoma.

Objectives: Limited data support the use of fasciotomies in acute limb ischemia (ALI) in patients with isolated arterial occlusion. This study describes an experience in which fasciotomies are not regularly performed post-revascularization. Methods: Using International Classification of Diseases, Ninth and Tenth Edition codes, patients presenting to the University of California Davis Medical Center between January 2003 and July 2018 with ALI, excluding those with traumatic injuries were identified. The primary outcome was major amputation, and the secondary outcome was foot drop. Additionally, the characteristics of those patients in each category of ischemic severity excluding those with grade 3 ischemia were summarized. Results: Of the 253 patients identified, revascularization was successful in 230 patients with 11 total fasciotomies performed. One hundred thirty-five patients were Rutherford Class 1/2A and 95 were 2B. In those with 1/2A ischemia, 134 (102 had >6 hours of symptoms) did not undergo fasciotomy with only one amputation occurring in this group. In those with 2B ischemia, 65 had >6 hours of symptoms; 58 did not undergo fasciotomy with 4 major amputations. In the 30 patients with ≤6 hours of ischemic symptoms, 27 did not undergo fasciotomy with 1 major amputation occurring in this group. There were no amputations in those patients who underwent fasciotomies. Additionally, there were 14 patients with a foot drop, of which 11 were in patients with 2B ischemia without fasciotomy. Conclusions: The data suggest that regardless of ischemic duration, 1/2A patients may not need fasciotomies, while those patients with 2B ischemia may benefit.

Retinitis Pigmentosa (RP) is a blinding disease that arises from loss of rods and subsequently cones. The P23H rhodopsin knock-in (P23H-KI) mouse develops retinal degeneration that mirrors RP phenotype in patients carrying the orthologous variant. Previously, we found that the P23H rhodopsin protein was degraded in P23H-KI retinas, and the Unfolded Protein Response (UPR) promoted P23H rhodopsin degradation in heterologous cells in vitro. Here, we investigated the role of a UPR regulator gene, activating transcription factor 6 (Atf6), in rhodopsin protein homeostasis in heterozygous P23H rhodopsin (Rho+/P23H) mice. Significantly increased rhodopsin protein levels were found in Atf6−/−Rho+/P23H retinas compared to Atf6+/−Rho+/P23H retinas at early ages (~ P12), while rhodopsin mRNA levels were not different. The IRE1 pathway of the UPR was hyper-activated in young Atf6−/−Rho+/P23H retinas, and photoreceptor layer thickness was unchanged at this early age in Rho+/P23H mice lacking Atf6. By contrast, older Atf6−/−Rho+/P23H mice developed significantly increased retinal degeneration in comparison to Atf6+/−Rho+/P23H mice in all retinal layers, accompanied by reduced rhodopsin protein levels. Our findings demonstrate that Atf6 is required for efficient clearance of rhodopsin protein in rod photoreceptors expressing P23H rhodopsin, and that loss of Atf6 ultimately accelerates retinal degeneration in P23H-KI mice.

Activating transcription factor 6 (ATF6), a key regulator of the unfolded protein response, is required for endoplasmic reticulum function and protein homeostasis. Variants of ATF6 that abrogate transcriptional activity cause morphologic and molecular defects in cones, manifesting clinically as the human vision loss disease achromatopsia (ACHM). ATF6 is expressed in all retinal cells. However, the effect of disease-associated ATF6 variants on other retinal cell types remains unclear. To investigate this question, we analyzed bulk RNA-sequencing transcriptomes from retinal organoids generated from patients with ACHM, carrying homozygous loss-of-function ATF6 variants. We identified marked dysregulation in mitochondrial respiratory complex gene expression and disrupted mitochondrial morphology in ACHM retinal organoids, indicating that loss of ATF6 leads to previously unappreciated mitochondrial defects in the retina. Next, we compared gene expression from control and ACHM retinal organoids with transcriptome profiles of seven major retinal cell types generated from recent single-cell transcriptomic maps of nondiseased human retina. Our analysis revealed pronounced down-regulation of cone genes and up-regulation in Müller glia genes, with no significant effects on other retinal cells. Overall, our analysis of ACHM patient retinal organoids identifies new cellular and molecular phenotypes in addition to cone dysfunction: activation of Müller cells, increased endoplasmic reticulum stress, disrupted mitochondrial structure, and elevated respiratory chain activity gene expression.

Sporadic Creutzfeldt-Jakob disease (sCJD) is the most commonly diagnosed human prion disease caused by the abnormal misfolding of the 'cellular' prion protein (PrP

An 84-year-old male with previously documented poor medical follow-up presented with progressive painless proptosis of the right eye. Right upper eyelid ptosis, limited motility, proptosis, and inferomedial displacement of the right globe were noted on the exam. Computed tomography (CT) imaging revealed a right retrobulbar extraconal heterogenous mass with ill-defined borders. Biopsy revealed a malignant adenocarcinoma with tumor markers suggestive of a colorectal primary. A rectal mass was identified during a systemic workup. After biopsy, the patient was diagnosed with stage IV metastatic rectal adenocarcinoma. He began palliative radiation therapy shortly following diagnosis.

Introduction: Uveal melanoma (UM) is the most common primary intraocular malignancy in adults, and despite treatment of the primary tumor, approximately 15%–50% of patients will develop metastatic disease. Based on gene expression profiling (GEPs), UM can be categorized as Class 1A (low metastatic risk), Class 1B (intermediate metastatic risk), or Class 2 (high metastatic risk). PReferentially expressed Antigen in MElanoma (PRAME) status is an independent prognostic UM biomarker and a potential target for immunotherapy in metastatic UM. PRAME expression status can be detected in tumors using reverse-transcription polymerase chain reaction (RT-PCR). More recently, immunohistochemistry (IHC) has been developed to detect PRAME protein expression. Here, we employed both techniques to evaluate PRAME expression in 18 UM enucleations. Methods: Tumor material from the 18 UM patients who underwent enucleation was collected by fine-needle aspiration before or during enucleation and sent for GEP and PRAME analysis by RT-PCR. Histologic sections from these patients were stained with an anti-PRAME monoclonal antibody. We collected patient demographics and tumor characteristics and included this with our analysis of GEP class, PRAME status by RT-PCR, and PRAME status by IHC. PRAME IHC and RT-PCR results were compared. Results: Twelve males (12/18) and 6 females (6/18) with an average age of 60.6 years underwent enucleation for UM. TNM staging of the UM diagnosed Stage I in 2 patients (2/18), Stage II in 7 patients (7/18), Stage III in 8 patients (8/18), and Stage IV in 1 (1/18). GEP was Class 1A in 6 tumors (6/18), Class 1B in 6 tumors (6/18), and Class 2 in 6 tumors (6/18). PRAME IHC showed diffusely positive labeling of all UM cells in 2/18 enucleations; negative IHC labeling of UM cells in 9/18 enucleations; and IHC labeling of subsets of UM cells in 7/18 enucleations. Eleven of the 17 UMs tested for PRAME by both RT-PCR and IHC had consistent PRAME results. In the remaining 6/17 cases tested by both modalities, PRAME results were discordant between RT-PCR and IHC. Conclusions: We find that PRAME IHC distinguishes PRAME-positive and PRAME-negative UM tumor cells. Interestingly, IHC reveals focal PRAME expression in subsets of tumor cells consistent with tumor heterogeneity. PRAME RT-PCR and IHC provide concordant results in most of our cases. We suggest that discordance in PRAME results could arise from spatial or temporal variation in PRAME expression between tumor cells. Further studies are required to determine the prognostic implications of PRAME IHC in UM.

Purpose To characterize the association of reticular pseudodrusen (RPD) with late-onset retinal degeneration (L-ORD) using multimodal imaging. Design Prospective, 2-center, longitudinal case series. Participants Twenty-nine patients with L-ORD. Methods All patients were evaluated within a 3-year interval with near-infrared reflectance, fundus autofluorescence, and spectral-domain OCT. In addition, a subset of patients also underwent indocyanine green angiography, fundus fluorescein angiography, mesopic microperimetry, and multifocal electroretinography. Main Outcome Measures Prevalence, topographic distribution, and temporal phenotypic changes of RPD in L-ORD. Results A total of 29 patients with molecularly confirmed L-ORD were included in this prospective study. Reticular pseudodrusen was detected in 18 patients (62%) at baseline, 10 of whom were men. The prevalence of RPD varied with age. The mean age of RPD patients was 57.3 ± 7.2 years. Reticular pseudodrusen was not seen in patients younger than the fifth decade of life (n = 3 patients) or in the eighth decade of life (n = 5 patients). Reticular pseudodrusen were found commonly in the macula with relative sparing of the fovea and also were identified in the peripheral retina. The morphologic features of RPD changed with follow-up. Two patients (3 eyes) demonstrated RPD regression. Conclusions Reticular pseudodrusen is found frequently in patients with L-ORD and at a younger age than in individuals with age-related macular degeneration (AMD). Reticular pseudodrusen exhibits quick formation and collapse, change in type and morphologic features with time, and relative foveal sparing and also has a peripheral retinal location in L-ORD.

Endoplasmic reticulum (ER) stress and Unfolded Protein Response (UPR) signaling promote the pathology of many human diseases. Loss-of-function variants of the UPR regulator Activating Transcription Factor 6 (ATF6) cause severe congenital vision loss diseases such as achromatopsia by unclear pathomechanisms. To investigate this, we generated retinal organoids from achromatopsia patient induced pluripotent stem cells carrying ATF6 disease variants and from gene-edited ATF6 null hESCs. We found that achromatopsia patient and ATF6 null retinal organoids failed to form cone structures concomitant with loss of cone phototransduction gene expression, while rod photoreceptors developed normally. Adaptive optics retinal imaging of achromatopsia patients carrying ATF6 variants also showed absence of cone inner/outer segment structures but preserved rod structures, mirroring the defect in cone formation observed in our retinal organoids. These results establish that ATF6 is essential for human cone development. Interestingly, we find that a selective small molecule ATF6 signaling agonist restores the transcriptional activity of some ATF6 disease-causing variants and stimulates cone growth and gene expression in patient retinal organoids carrying these variants. These findings support that pharmacologic targeting of the ATF6 pathway can promote human cone development and should be further explored for blinding retinal diseases.

Radiation-induced malignancy is rare, occurring in approximately 0.4%-1.0% of patients receiving external beam radiation therapy. Sarcomas and squamous cell carcinomas are among the most common types of cancers to occur. A 74-year-old woman presented with redness and swelling in the right periorbital region. She had history of multiple recurrent ameloblastoma of the right maxilla, invading the right orbital floor status post 4 surgical resections and 66 Gray external beam radiotherapy 5 years prior. MRI showed a poorly circumscribed mass involving the inferior and lateral orbit. Orbital biopsy revealed clear cell carcinoma with hyalinizing sclerosis and Ewing sarcoma breakpoint region 1 gene arrangement. Due to the extent of orbital disease and presence of perineural invasion, she underwent orbital exenteration. Hyalinizing clear cell carcinoma, a rare cancer, has not been reported to occur in the orbit following radiation. This case highlights the importance of lifetime monitoring in patients who have undergone radiation therapy.

G protein mutations are common in uveal melanomas, and the vast majority target amino acid residue Q209 in either GNAQ or GNA11. The GNAQ R183Q mutation is found in a small fraction of uveal melanomas. We report a patient with an unusual presentation of uveal melanoma arising at an early age in the setting of congenital skin and ocular surface melanosis. A 34-year-old Hispanic female with congenital bilateral nevus of Ota and ocular surface melanosis presented with progressive loss of visual acuity and was found to have a juxtapapillary uveal melanoma. She was treated with brachytherapy, but the tumor relapsed. She underwent enucleation that revealed mixed spindle and epithelioid uveal melanoma cells with no extraocular or lymphovascular spread. Next-generation sequencing performed on DNA isolated from the enucleation specimen identified a GNAQ R183Q mutation and a PMS1 truncation mutation. Cytogenetic profiling revealed no monosomy 3. These findings raise the possibility that uveal melanomas bearing G protein R183 mutations may have distinct clinicopathologic profiles compared to those with Q209 mutations. Furthermore, this is the first reported case of a mutation in the mismatch repair gene PMS1 associated with uveal melanoma.

The coupling of large telescopes to astronomical instruments has historically been challenging due to the tension between instrument throughput and stability. Light from the telescope can either be injected wholesale into the instrument, maintaining high throughput at the cost of point-spread function (PSF) stability, or the time-varying components of the light can be filtered out with single-mode fibers (SMFs), maintaining instrument stability at the cost of light loss. Today, the field of astrophotonics provides a potential resolution to the throughput-stability tension in the form of the photonic lantern (PL): a tapered waveguide that can couple a time-varying and aberrated PSF into multiple diffraction-limited beams at an efficiency that greatly surpasses direct SMF injection. As a result, lantern-fed instruments retain the stability of SMF-fed instruments while increasing their throughput. To this end, we present a series of numerical simulations characterizing PL performance as a function of lantern geometry, wavelength, and wavefront error (WFE), aimed at guiding the design of future diffraction-limited spectrometers. These characterizations include a first look at the interaction between PLs and phase-induced amplitude apodization (PIAA) optics. We find that Gaussian-mapping beam-shaping optics can enhance coupling into 3-port lanterns but offer diminishing gains with larger lanterns. In the y and J band (0.97–1.35 µm) region, with moderately high WFE ( ∼ 10 % Strehl ratio), a 3-port lantern in conjunction with beam-shaping optics strikes a good balance between pixel count and throughput gains. If pixels are not a constraint, and the flux in each port will be dominated by photon noise, then larger port count lanterns will provide further coupling gains due to a greater resilience to tip-tilt errors. Finally, we show that lanterns can maintain high operating efficiencies over large wavelength bands where the number of guided modes at the lantern entrance drops, if care is taken to minimize the attenuation of weakly radiative input modes.

Purpose To investigate the use of an amyloid-targeting fluorescent probe, ARCAM-1, to identify amyloid-containing deposits in the retina of a transgenic mouse model of Alzheimer's disease (AD) and in human postmortem AD patients. Methods Aged APP/PS1 transgenic AD and wild-type (WT) mice were given an intraperitoneal (IP) injection of ARCAM-1 and their retinas imaged in vivo using a fluorescence ophthalmoscope. Eyes were enucleated and dissected for ex vivo inspection of retinal amyloid deposits. Additionally, formalin-fixed eyes from human AD and control patients were dissected, and the retinas were stained using ARCAM-1 or with an anti-amyloid-β antibody. Confocal microscopy was used to image amyloid-containing deposits stained with ARCAM-1 or with immunostaining. Results Four out of eight APP/PS1 mice showed the presence of amyloid aggregates in the retina during antemortem imaging. Retinas from three human AD patients stained with ARCAM-1 showed an apparent increased density of fluorescently labeled amyloid-containing deposits compared to the retinas from two healthy, cognitively normal (CN) patients. Immunolabeling confirmed the presence of amyloid deposits in both the retinal neuronal layers and in retinal vasculature. Conclusions ARCAM-1 facilitates antemortem detection of amyloid aggregates in the retina of a mouse model for AD, and postmortem detection of amyloid-containing deposits in human retinal tissues from AD patients. These results support the hypothesis of AD pathology manifesting in the eye and highlight a novel area for fluorophore development for the optical detection of retinal amyloid in AD patients. Translational relevance This paper represents an initial examination for potential translation of an amyloid-targeting fluorescent probe to a retinal imaging agent for aiding in the diagnosis of Alzheimer's disease.

Aim: Nonalcoholic fatty liver disease (NAFLD)-associated hepatocellular carcinoma (HCC) is projected to become the leading indication for liver transplantation. Previous studies indicate that tumor growth rates (TGR) may predict survival and were helpful in determining HCC surveillance intervals. Therefore, we aimed to determine its usefulness in predicting clinical outcomes and treatments. Methods: We conducted a retrospective study of hepatitis B, C and NAFLD-HCC cases. TGR was measured using 2-consecutive pre-treatment contrast-enhanced imaging studies ≥ 25 days apart. A multivariate regression model was used to determine predictors of TGR. In addition, the Cox regression model was used to evaluate the relationship between TGR and overall survival. Results: From 2000–2019, the study cohort comprised 38, 60, and 47 HBV, HCV, and NAFLD patients, respectively, with TGRs. NAFLD-HCC tumor size was inversely correlated to the extent of liver disease as measured by Child-Pugh score (7.2 cm in non-cirrhosis; 3.7 cm, 2.6 cm, and 2.1 cm in Child A, B, and C, respectively; P < 0.001). After adjusting for baseline characteristics, the TGR per month was fastest in HBV (9.4%, 95%CI: 6.3%-12.5%) compared to HCV (4.9%, 95%CI: 2.8%-7%) and NAFLD patients (3.6%, 95%CI: 1.6%-6.7%). Predictors of TGR included elevated AFP, low albumin, and smaller tumor size. Fast TGR in viral etiologies had higher mortality [adj. hazard ratio (HR) = 2.6, 95%CI: 1.2–5.7, P = 0.02] than slow TGRs, independent of treatments. Fast TGR in NAFLD had a trend towards higher mortality (HR = 3.6, 95%CI: 0.95–13.3, P = 0.059). Conclusion: NAFLD-HCC patients have more indolent growths than viral-related HCC TGRs. The addition of TGR as a biomarker may assist in stratifying treatment options.

Dysregulation of the endoplasmic reticulum (ER) Unfolded Protein Response (UPR) is implicated in the pathology of many human diseases associated with ER stress. Inactivating genetic variants in the UPR regulator Activating Transcription Factor 6 (ATF6) cause severe congenital heritable vision loss in patients by an unknown pathomechanism. To investigate this, we generated retinal organoids from patient iPSCs carrying ATF6 disease-causing variants and ATF6 null hESCs generated by CRISPR. Interestingly, we found that cone photoreceptor cells in ATF6 mutant retinal organoids lacked inner and outer segments concomitant with absence of cone phototransduction gene expression; while rod photoreceptors developed normally. Adaptive optics retinal imaging of patients with disease-causing variants in ATF6 also showed absence of cone inner/outer segment structures but preserved rod structures, mirroring the phenotypes observed in our retinal organoids. These results reveal that ATF6 is essential for the formation of human cone photoreceptors, and associated absence of cone phototransduction components explains the severe visual impairment in patients with ATF6 -associated retinopathy. Moreover, we show that a selective small molecule ATF6 activator compound restores the transcriptional activity of ATF6 disease-causing variants and stimulates the growth of cone photoreceptors in patient retinal organoids, demonstrating that pharmacologic targeting of ATF6 signaling is a therapeutic strategy that needs to be further explored for blinding retinal diseases.

The authors report an unusual case of lung adenocarcinoma metastasis to the lacrimal sac. A 61-year-old woman with stage IV non-small cell lung cancer presented with left facial pain and epiphora. She was found to have an elevated tear meniscus associated with a firm, fixed medial canthal mass. Orbital imaging demonstrated nodular enlargement of the lacrimal drainage apparatus. Biopsy of the lacrimal sac was performed, and it revealed a metastatic lung adenocarcinoma. The patient received targeted radiation therapy to the lacrimal sac, and her dose of maintenance chemotherapy was increased. The patient's symptoms have since improved. This case of lung cancer involving the lacrimal sac highlights the importance of thorough oncologic surveillance, even with respect to locations atypical for metastatic spread.

Worldwide, nonalcoholic fatty liver disease (NAFLD) has reached epidemic proportions and in parallel, hepatocellular carcinoma (HCC) has become one of the fastest growing cancers. Epidemiological studies have not only shed light on the prevalence and incidence of the disease but have also unmasked important environmental risk factors, including the role of diabetes and dyslipidemia in disease pathogenesis. Genetic association studies have identified single nucleotide polymorphisms implicated in NAFLD-HCC, many of which are part of lipid metabolism pathways. Through these clinical studies and subsequently, translational and basic research, the role of statins as a chemoprotective agent has also emerged with ongoing clinical trials assessing their utility in HCC prevention and treatment. In this review, we summarize the recent epidemiological studies describing the burden of NAFLD-HCC in different patient populations and countries. We discuss the genetic and environmental risk factors for NAFLD-HCC and highlight the chemoprotective role of statins and aspirin. We also summarize what is known about NAFLD-HCC in the cirrhosis and non-cirrhosis populations and briefly address the role of surveillance in NAFLD-HCC patients.

Hepatocellular carcinoma (HCC) is a leading cause of cancer‐related death worldwide. Society guidelines recommend surveillance with abdominal ultrasound with or without serum alpha‐fetoprotein every 6 months for adults at increased risk of developing HCC. However, adherence is often suboptimal. We assessed the feasibility of a coordinated telephone outreach program for unscreened patients with cirrhosis within the Veteran’s Affairs (VA) health care system. Using a patient care dashboard of advanced chronic liver disease in the VA Greater Los Angeles Healthcare System, we identified veterans with a diagnosis of cirrhosis, a platelet count ≤ 150,000/uL, and no documented HCC surveillance in the previous 8 months. Eligible veterans received a telephone call from a patient navigator to describe the risks and benefits of HCC surveillance. Orders for an abdominal ultrasound and alpha‐fetoprotein were placed for veterans who agreed to surveillance. Veterans who were not reached by telephone received an informational letter by mail to encourage participation. Of the 129 veterans who met the eligibility criteria, most were male (96.9%). The most common etiology for cirrhosis was hepatitis C (64.3%), and most of the patients had compensated cirrhosis (68.2%). The patient navigators reached 32.5% of patients by phone. Patients in each group were similar across clinical and demographic characteristics. Patients who were called were more likely to undergo surveillance (adjusted odds ratio = 2.56, 95% confidence interval: 1.03‐6.33). Most of the patients (72.1%) completed abdominal imaging when reached by phone. Conclusion: Targeted outreach increased uptake of HCC surveillance among patients with cirrhosis in a large, integrated, VA health care system., HCC surveillance in patients with cirrhosis is recommended by society guidelines, yet suboptimal. In this quality improvement outreach pilot study within a large VA, we demonstrate that HCC surveillance improves by telephone and mail outreach of patients who have confirmed cirrhosis and had not been screened.

Achromatopsia (ACHM) is an autosomal recessive disease that results in severe visual loss. Symptoms of ACHM include impaired visual acuity, nystagmus, and photoaversion starting from infancy; furthermore, ACHM is associated with bilateral foveal hypoplasia and absent or severely reduced cone photoreceptor function on electroretinography. Here, we performed genetic sequencing in 3 patients from 2 families with ACHM, identifying and functionally characterizing 2 mutations in the activating transcription factor 6 (ATF6) gene. We identified a homozygous deletion covering exons 8-14 of the ATF6 gene from 2 siblings from the same family. In another patient from a different family, we identified a heterozygous deletion covering exons 2 and 3 of the ATF6 gene found in trans with a previously identified ATF6 c.970C>T (p.Arg324Cys) ACHM disease allele. Recombinant ATF6 proteins bearing these exon deletions showed markedly impaired transcriptional activity by qPCR and RNA-Seq analysis compared with WT-ATF6. Finally, RNAscope revealed that ATF6 and the related ATF6B transcripts were expressed in cones as well as in all retinal layers in normal human retina. Overall, our data identify loss-of-function ATF6 disease alleles that cause human foveal disease.

Purpose Uveal melanomas are associated with characteristic genetic changes. Germline mutations in mismatch repair (MMR) genes and microsatellite instability have been implicated in the development of numerous malignant neoplasms such as colon and ovarian cancers. The frequency of MMR defects in uveal melanomas has yet to be determined. Methods Here, we analyzed the frequency of MMR gene mutations in uveal melanoma specimens from the University of California, San Diego (UCSD), The Cancer Genome Atlas (TGCA), and the Catalogue of Somatic Mutations in Cancer (COSMIC). Results We identified only two mutations in a MMR gene: one premature stop codon in the PMS gene within the UCSD cohort (0.5% frequency) and one in-frame deletion in MSH3 within the COSMIC database (0.8% frequency). We report copy number variation of MLH1 in monosomy 3 and show decreased mRNA expression of MLH1 in uveal melanoma specimens with monosomy 3. Expression levels of MLH1 were not found to correlate with the observed number of total mutations. Conclusion Overall, we show that mutations in MMR genes in uveal melanoma specimens are exceedingly rare, and although one copy of MLH1 is lost in monosomy 3, it does not seem to have pathologic consequences in uveal melanoma pathogenesis.

ObjectiveStudies evaluating major amputation after initial minor amputation are few with rates of subsequent major amputation ranging from 14% to 35% with limited understanding of associated comorbidities and time to limb loss. The aim of this study is to determine the major amputation rates for patients who had already undergone an initial minor amputation and determine which factors are associated with the need for subsequent major amputation.MethodsUsing statewide data between 2005 and 2013, patients with peripheral artery disease (PAD), diabetes mellitus (DM), and combined PAD/DM who had a lower extremity ulcer and who had also undergone a minor amputation were identified. These patients were evaluated for the rate of subsequent major amputation and competing risk Cox proportional hazards modeling was used to study which factors were associated with the risk of subsequent limb loss.ResultsThe cohort consisted of 11,597 patients (DM, n= 4254; PAD, n= 2142; PAD/DM, n= 5201) with lower extremity ulcers who underwent an initial minor amputation. The rate of any subsequent amputation was highest in patients with PAD/DM (23% vs DM= 17%, PAD= 17%; P= not statistically significant). The rate of subsequent minor amputation was 16% in the PAD/DM versus 15.2% in PAD and 12.2% in patients with DM (P< .001). Patients with PAD/DM had the highest rate of subsequent major amputation (6.3% vs DM= 5.2%, PAD= 2.1%; P< .001). There was no statistically significant difference in the median time to major amputation among the three groups (PAD/DM, 13months; DM, 14months; PAD, 8.6months; P= NS). Patients who were revascularized before a repeat minor amputation had a decreased risk of a major amputation compared with those who were intervened on after a repeat minor amputation (hazard ratio, 0.002; 95% confidence interval, 0-0.22). Patients treated completely in the outpatient setting were also less likely to undergo subsequent major amputation (hazard ratio, 0.7; 95% confidence interval, 0.5-0.98) compared with those who required hospitalization or presented to the emergency room.ConclusionsPatients with ulcers and combined PAD and DM have a higher risk for secondary major and minor amputation than patients with either disease alone with half of the limb loss occurring at approximately 1year after the initial minor amputation. Additionally, early diagnosis and appropriate referral may result in decreased limb loss for these patients.

Dysregulation of the endoplasmic reticulum (ER) Unfolded Protein Response (UPR) is implicated in the pathology of many human diseases associated with ER stress. Inactivating genetic variants in the UPR regulator Activating Transcription Factor 6 (ATF6) cause severe congenital heritable vision loss in patients by an unknown pathomechanism. To investigate this, we generated retinal organoids from patient iPSCs carrying ATF6 disease-causing variants and ATF6 null hESCs generated by CRISPR. Interestingly, we found that cone photoreceptor cells in ATF6 mutant retinal organoids lacked inner and outer segments concomitant with absence of cone phototransduction gene expression; while rod photoreceptors developed normally. Adaptive optics retinal imaging of patients with disease-causing variants in ATF6 also showed absence of cone inner/outer segment structures but preserved rod structures, mirroring the phenotypes observed in our retinal organoids. These results reveal that ATF6 is essential for the formation of human cone photoreceptors, and associated absence of cone phototransduction components explains the severe visual impairment in patients with ATF6 -associated retinopathy. Moreover, we show that a selective small molecule ATF6 activator compound restores the transcriptional activity of ATF6 disease-causing variants and stimulates the growth of cone photoreceptors in patient retinal organoids, demonstrating that pharmacologic targeting of ATF6 signaling is a therapeutic strategy that needs to be further explored for blinding retinal diseases.

The human eye is the organ that is able to react to light in order to provide sharp three-dimensional and colored images. Unfortunately, the health of the eye can be impacted by various stimuli that can lead to vision loss, such as environmental changes, genetic mutations, or aging. Endoplasmic reticulum (ER) stress and unfolded protein response (UPR) signaling have been detected in many diverse ocular diseases, and chemical and genetic approaches to modulate ER stress and specific UPR regulatory molecules have shown beneficial effects in animal models of eye disease. This review highlights specific eye diseases associated with ER stress and UPR activity, based on a recent symposia exploring this theme.

We present the case of a patient with a sinonasal mass diagnosed as glomangiopericytoma on initial resection. The patient experienced multiple recurrences over ten years, undergoing endoscopic resection each time and ultimately adjunctive radiation therapy. Cytogenetic testing of the fourth recurrence was positive for FUS-ERG fusion, a genetic abnormality seen in a rare subset of Ewing tumors and the first case to be described in a sinonasal tumor. Given the emergence of new immunophenotyping, clinicians should exercise a low threshold for cytogenetic testing in cases where a tumor exhibits characteristics atypical of the initial pathological diagnosis.

High dose ascorbic acid (HDAA) has been touted to ameliorate inflammation and reduce fluid requirements during burn shock resuscitation (BSR). Whether this leads to improved outcomes is not known. The authors' aim for this study was to compare ventilator days, ventilator-associated pneumonia, and mortality between patients who did and did not receive HDAA during BSR.The authors performed a retrospective case control study from 2012 to 2015. They identified 38 patients (HDAA) who received HDAA during BSR. Using age and %TBSA, the authors identified and matched 42 control patients (CTL) who did not receive HDAA for BSR during that same time period. The authors collected data for age, %TBSA, hospital days (LOS), ventilator days (VENT), inhalation injury (INH), ventilator-associated pneumonia (VAP), and mortality (MORT).There were no differences in age and %TBSA or %TBSA of third-degree burn injury between groups. There was no significant difference in the incidence of INH (HDAA-52% vs CTL-36%, P = .17) and the groups had similar LOS and VENT. Additionally, there was no significant difference in VAP incidence (HDAA-29% vs CTL-14%, P = .13) or mortality (HDAA-26% vs CTL-23%, P = .8). HDAA patients had a numerically higher incidence of acute renal failure requiring dialysis (23 vs 7%, P = .06) which was confirmed in a multivariate analysis (odds ratio 5.4; 95% confidence interval 1.1-26). HDAA, while potentially reducing inflammation and fluid requirements during BSR, may not improve any meaningful outcomes such as ventilator requirements, ventilator-associated pneumonia, and mortality.

Inherited retinal dystrophies (IRDs) are clinically and genetically highly heterogeneous, making clinical diagnosis difficult. The advances in high-throughput sequencing (ie, panel, exome and genome sequencing) have proven highly effective on defining the molecular basis of these disorders by identifying the underlying variants in the respective gene. Here we report two siblings affected by an IRD phenotype and a novel homozygous c.1691A>G (p.(Asp564Gly)) ATF6 (activating transcription factor 6A) missense substitution identified by whole exome sequencing analysis. The pathogenicity of the variant was confirmed by functional analyses done on patients’ fibroblasts and on recombinant p.(Asp564Gly) protein. The ATF6Asp564Gly/Asp564Gly variant shows impaired production of the ATF6 cleaved transcriptional activator domain in response to endoplasmic reticulum stress. Detailed phenotypic examination revealed extinguished cone responses but also decreased rod responses together with the ability to discriminate some colours suggestive rather for cone-rod dystrophy than achromatopsia.

Activating transcription factor 6 (ATF6) is a key regulator of the unfolded protein response (UPR). In response to endoplasmic reticulum (ER) stress, ATF6 is transported from the ER to the Golgi apparatus where it is cleaved by intramembrane proteolysis, releasing its cytosolic fragment. The cleaved ATF6 fragment, which is a basic leucine zipper (bZip) transcription factor, translocates to the nucleus and upregulates the expression of ER protein-folding chaperones and enzymes. Mutations in ATF6 cause heritable forms of cone photoreceptor dysfunction diseases. These mutations include missense, nonsense, splice site, and deletion or duplication changes found across the entire ATF6. To date, there are 11 ATF6 mutations reported, and we classified them into three classes based on their functional defects that interrupt distinct steps in the ATF6 signaling pathway.

Vehicular ad hoc networks (VANETs) are designed to provide traffic safety by enabling vehicles to broadcast information-such as speed, location and heading-through inter-vehicular communications to proactively avoid collisions. However, the attacks targeting these networks might overshadow their advantages if not protected against. One powerful threat against VANETs is vehicular botnets. In our earlier work, we demonstrated several vehicular botnet attacks that can have damaging impacts on the security and privacy of VANETs. In this paper, we present SHIELDNET, the first detection mechanism against vehicular botnets. Similar to the detection approaches against Internet botnets, we target the vehicular botnet communication and use several machine learning techniques to identify vehicular bots. We show via simulation that SHIELDNET can identify 77 percent of the vehicular bots. We propose several improvements on the VANET standards and show that their existing vulnerabilities make an effective defense against vehicular botnets infeasible.

Background: p16 immunohistochemistry is widely used to diagnose human papillomavirus (HPV)-related squamous neoplasms of cervix, anogenital, head, and neck tissues. The incidence of these HPV-related squamous neoplasms is markedly increased in the HIV-infected population. Ocular surface squamous neoplasia (OSSN) is also more common in HIV-infected patients. However, the expression pattern of p16 in OSSN among HIV-infected patients is unclear. Here, we examined the expression of p16 in OSSN surgical excisions collected from a large HIV-infected cohort from ­Mozambique. Methods: OSSN surgical tissue specimens were collected from 75 Mozambican patients. Formalin-fixed, paraffin-embedded tissue blocks from these OSSNs were sectioned, stained with hematoxylin and eosin (H&E), and p16 expression by immunohistochemistry. H&E slides were reviewed to determine if OSSNs were noninvasive conjunctival intraepithelial neoplasms or invasive squamous cell carcinomas (SCC). Cases were classified as p16 positive or negative based on diffuse nuclear and cytoplasmic expression of p16 in neoplastic cells. Results: p16 positivity was found in a minority of OSSN cases (14/75). p16 positivity was significantly associated with the invasive SCC type of OSSN in HIV-infected patients (p value of 0.026). Conclusions: The majority of OSSNs in our HIV-infected cohort do not express p16. However, those cases that are p16-positive are significantly more likely to be the invasive SCC form of OSSN. We propose that p16 expression may identify more aggressive OSSNs in HIV-infected populations.

Background: Critical limb ischemia remains a difficult disease to treat, with limited level one data. The BEST-CLI trial (Best Endovascular vs Best Open Surgical Therapy in Patients with Critical Limb Ischemia) is attempting to answer whether initial treatment with open surgical bypass or endovascular therapy improves outcomes, although it remains in enrollment. This study aims to compare amputation-free survival and reintervention rates in patients treated with initial open surgical bypass or endovascular intervention for ischemic ulcers of the lower extremities. Methods and Results: Using California nonfederal hospital data linked to statewide death data, all patients with lower extremity ulcers and a diagnosis of peripheral artery disease who underwent a revascularization procedure from 2005 to 2013 were identified. Propensity scores were formulated from baseline patient characteristics. Inverse probability weighting was used with Kaplan–Meier analysis to determine amputation-free survival and time to reintervention for open versus endovascular treatment. Mixed-effects Cox proportional hazards modeling was used to adjust for patient ability to manage their disease and hospital revascularization volume. A total of 16 800 patients were identified. Open surgical bypass was the initial treatment in 5970 (36%) while 10 830 (64%) underwent endovascular interventions. Patients in the endovascular group were slightly younger compared with the open group (70 versus 71 years, ±12 years; P P values Conclusions: Patients with critical limb ischemia have multiple comorbidities, and initial surgical bypass is associated with poorer amputation-free survival compared with an endovascular-first approach, perhaps due to increased severity of wounds at the time of presentation.

Endoplasmic reticulum (ER) stress activates the unfolded protein response (UPR), which reduces levels of misfolded proteins. However, if ER homeostasis is not restored and the UPR remains chronically activated, cells undergo apoptosis. The UPR regulator, PKR-like endoplasmic reticulum kinase (PERK), plays an important role in promoting cell death when persistently activated; however, the underlying mechanisms are poorly understood. Here, we profiled the microRNA (miRNA) transcriptome in human cells exposed to ER stress and identified miRNAs that are selectively induced by PERK signaling. We found that expression of a PERK-induced miRNA, miR-483, promotes apoptosis in human cells. miR-483 induction was mediated by a transcription factor downstream of PERK, activating transcription factor 4 (ATF4), but not by the CHOP transcription factor. We identified the creatine kinase brain-type (CKB) gene, encoding an enzyme that maintains cellular ATP reserves through phosphocreatine production, as being repressed during the UPR and targeted by miR-483. We found that ER stress, selective PERK activation, and CKB knockdown all decrease cellular ATP levels, leading to increased vulnerability to ER stress-induced cell death. Our findings identify miR-483 as a downstream target of the PERK branch of the UPR. We propose that disruption of cellular ATP homeostasis through miR-483-mediated CKB silencing promotes ER stress-induced apoptosis.

A 5-year-old otherwise healthy girl presented to the oculoplastic service with a painless superotemporal subconjunctival mass in the left eye. Visual acuity was within normal limits, and there was no evidence of proptosis or orbital enlargement. Excision was performed to remove the anterior portion of the mass for alleviation of symptoms. On histopathological analysis, the mass was comprised of fibroadipose tissue consistent with dermolipoma and contained a hard nodule found to be a calcified tooth. In the periocular region, odontogenic choristoma (tooth) is a rare lesion, and has been reported to occur within teratomas, dermoid cysts, and displaced oral embryonic epithelium. We describe an unusual case of a tooth occurring within a sporadic dermolipoma. The clinical presentation, examination, management, and histopathology are reviewed.

Photoreceptors are specialized sensory neurons essential for light detection in the human eye. Photoreceptor cell dysfunction and death cause vision loss in many eye diseases such as retinitis pigmentosa and achromatopsia. Endoplasmic reticulum (ER) stress and Unfolded Protein Response (UPR) signaling have been implicated in the development and pathology of heritable forms of retinitis pigmentosa and achromatopsia. We review the role of ER stress and UPR in retinitis pigmentosa arising from misfolded rhodopsins (RHO) and in achromatopsia arising from genetic mutations in Activating Transcription Factor 6 (ATF6). This article is part of a Special Issue entitled SI:ER stress.

Two patients, previously diagnosed and treated for euthyroid, autoantibody-negative thyroid eye disease, presented with active orbitopathy. An atypical disease course and presentation prompted orbital biopsy. Extraocular muscle histopathology demonstrated noncaseating granulomatous inflammation consistent with presumed orbital sarcoidosis involving multiple extraocular muscles, including the inferior oblique in one of the cases. These two cases emphasize the importance of a broad differential diagnosis and the utility of an orbital biopsy in the context of an unusual disease presentation or poor treatment response. The patients’ clinical course is discussed alongside important clinical signs, imaging findings, and biopsy results that support a diagnosis of isolated orbital sarcoidosis.

In the tumor microenvironment, local immune dysregulation is driven in part by macrophages and dendritic cells that are polarized to a mixed proinflammatory/immune-suppressive phenotype. The unfolded protein response (UPR) is emerging as the possible origin of these events. Here we report that the inositol-requiring enzyme 1 (IRE1α) branch of the UPR is directly involved in the polarization of macrophages in vitro and in vivo, including the up-regulation of interleukin 6 (IL-6), IL-23, Arginase1, as well as surface expression of CD86 and programmed death ligand 1 (PD-L1). Macrophages in which the IRE1α/X-box binding protein 1 (Xbp1) axis is blocked pharmacologically or deleted genetically have significantly reduced polarization and CD86 and PD-L1 expression, which was induced independent of IFNγ signaling, suggesting a novel mechanism in PD-L1 regulation in macrophages. Mice with IRE1α- but not Xbp1-deficient macrophages showed greater survival than controls when implanted with B16.F10 melanoma cells. Remarkably, we found a significant association between the IRE1α gene signature and CD274 gene expression in tumor-infiltrating macrophages in humans. RNA sequencing (RNASeq) analysis showed that bone marrow–derived macrophages with IRE1α deletion lose the integrity of the gene connectivity characteristic of regulated IRE1α-dependent decay (RIDD) and the ability to activate CD274 gene expression. Thus, the IRE1α/Xbp1 axis drives the polarization of macrophages in the tumor microenvironment initiating a complex immune dysregulation leading to failure of local immune surveillance., Tumor-infiltrating macrophages possess characteristics of an active unfolded protein response (UPR) associated with a mixed proinflammatory/immune-suppressive phenotype. This study shows that these characteristics are driven by the IRE1a/Xbp1 axis of the UPR, suggesting that targeting the UPR may increase the response to immune checkpoint blockade therapy.

Switch/sucrose non-fermentable-related matrix-associated actin-dependent regulator of chromatin subfamily B member 1 (SMARCB1), also known as integrase interactor 1-deficient sinonasal carcinoma, is a rare entity that was first described in 2014. Since then, there have been 39 cases published in the literature, with basaloid or plasmacytoid/rhabdoid morphology being the most common pathological subtype. We report a patient with switch/sucrose non-fermentable-related matrix-associated actin-dependent regulator of chromatin subfamily B member 1 (integrase interactor 1)-deficient sinonasal carcinoma who had permanent vision loss after valsalva-induced acute hemorrhage and resultant orbital compartment syndrome.