Management of Mammographic Architectural Distortion Based on Contrast-enhanced MRI and US Correlation.

OBJECTIVE: The objective was to evaluate outcomes of mammographic architectural distortion (AD) with and without MRI and US correlates. METHODS: A retrospective review of unexplained mammographic AD with subsequent MRI from January 1, 2007 to September 30, 2017 was performed using a reader-based study design. Mammographic, MRI, and US features and outcomes were documented. Truth was based on biopsy results or minimum two-year imaging follow-up. Measures of diagnostic accuracy were calculated. RESULTS: Fifty-six cases of AD were included: 29 (51.8%) detected on 2D mammogram and 27 (48.2%) detected on digital breast tomosynthesis. Of 35.7% (20/56) with MRI correlate, 40.0% (8/20) were enhancing masses, 55.0% (11/20) were non-mass enhancement (NME), and 5.0% (1/20) were nonenhancing AD. Of eight enhancing masses, 75.0% (6/8) were invasive cancers, and 25.0% (2/8) were high-risk lesions. Of 11 NME, 18.2% (2/11) were ductal carcinoma in situ, 36.4% (4/11) were high-risk lesions, and 45.4% (5/11) were benign. Of 64.3% (36/56) without MRI correlate, 94.4% (34/36) were benign by pathology or follow-up, one (2.8%, 1/36) was a 4-mm focus of invasive cancer with US correlate, and one (1/36, 2.8%) was a high-risk lesion. Of cases without MRI and US correlates, one (3.0%, 1/33) was a high-risk lesion and 97.0% (32/33) were benign. The negative predictive value of mammographic AD without MRI correlate was 97.2% (35/36) and without both MRI and US correlates was 100.0% (33/33). CONCLUSION: Mammographic AD without MRI or US correlate was not cancer in our small cohort and follow-up could be considered, reducing interventions.

Environmental oxygen affects ex vivo growth and proliferation of mesenchymal progenitors by modulating mitogen-activated protein kinase and mammalian target of rapamycin signaling.

BACKGROUND AIMS: Stem and progenitor cells of hematopoietic and mesenchymal lineages reside in the bone marrow under low oxygen (O2) saturation. O2 levels used in ex vivo expansion of multipotent mesenchymal stromal cells (MSCs) affect proliferation, metabolism and differentiation. METHODS: Using cell-based assays and transcriptome and proteome data, the authors compared MSC cultures simultaneously grown under a conventional 19.95% O2 atmosphere or at 5% O2. RESULTS: In 5% O2, MSCs showed better proliferation and higher self-renewal ability, most probably sustained by enhanced signaling activity of mitogen-activated protein kinase and mammalian target of rapamycin pathways. Non-oxidative glycolysis-based energy metabolism supported growth and proliferation in 5% O2 cultures, whereas MSCs grown under 19.95% O2 also utilized oxidative phosphorylation. Cytoprotection mechanisms used by cells under 5% O2 differed from 19.95% O2  suggesting differences in the triggers of cell stress between these two O2  conditions. CONCLUSIONS: Based on the potential benefits for the growth and metabolism of MSCs, the authors propose the use of 5% O2 for MSC culture.

Helicobacter pylori Infection and Non-alcoholic Fatty Liver Disease: A Systematic Review and Meta-Analysis.

Since numerous studies have stated that there may be a relationship between Helicobacter pylori infection and nonalcoholic fatty liver disease, and because of the high prevalence of both conditions worldwide, this study investigated the risk of non-alcoholic fatty liver disease in patients infected with H. pylori. Following a systematic review of PubMed, Scopus, Web of Science and Embase, and a search in Google Scholar using MeSH terms such as H. pylori and non-alcoholic fatty liver disease, the relevant papers up to November 2020 were reviewed. All cohort, case-control, and cross-sectional studies that examined the risk of developing non-alcoholic fatty liver disease in patients infected with H. pylori entered this study. A meta-analysis was conducted in STATA 11. This systematic review examined 22 papers with 117 117 participants (33 711 patients infected with H. pylori and 83 406 participants as control) and 20 studies were subjected to meta-analysis The results indicated a 22% to 27% increase in the risk of developing non-alcoholic fatty liver disease in patients infected with H. pylori (crude odds ratio: 1.27, 95% CI: 1.17-1.33; and adjusted odds ratio: 1.22, 95% CI: 1.09-1.35). According to the subgroup analysis, the study region, sample size, and the method of diagnosing H. pylori were the factors contributing to the high heterogeneity. The meta-analysis revealed the increased risk of developing non-alcoholic fatty liver disease in patients infected with H. pylori. This indicates that H. pylori is a serious risk factor in patients susceptible to NAFLD.

Factors Associated With Background Parenchymal Enhancement on Contrast-Enhanced Mammography.

OBJECTIVE. The purpose of this study was to determine the relationship between background parenchymal enhancement (BPE) on contrast-enhanced mammography (CEM) and breast tissue density, menstrual status, endocrine therapy, and risk factors for breast cancer and also to evaluate interreader agreement on classification of BPE on CEM. MATERIALS AND METHODS. Five subspecialty-trained breast radiologists independently and blindly graded tissue density (with fatty tissue and scattered fibroglandular tissue classified as nondense tissue and with heterogeneously dense and extremely dense classified as dense tissue) and BPE (with minimal or mild BPE categorized as low BPE and moderate or marked BPE categorized as high BPE) on CEM examinations performed from 2014 to 2018. Electronic medical charts were reviewed for information on menstrual status, endocrine therapy, history of breast surgery, and other risk factors for breast cancer. Comparisons were performed using the Kruskal-Wallis test, Mann-Whitney test, and Spearman rank correlation. Interreader agreement was estimated using the Fleiss kappa test. RESULTS. A total of 202 patients (mean [± SD] age, 54 ± 10 years; range, 25-84 years) underwent CEM. Tissue density was categorized as fatty in two patients (1%), scattered fibroglandular in 67 patients (33%), heterogeneously dense in 117 patients (58%), and extremely dense in 16 patients (8%). Among the 202 patients, BPE was minimal in 77 (38%), mild in 80 (40%), moderate in 31 (15%), and marked in 14 (7%). Dense breasts, younger age, premenopausal status, no history of endocrine therapy, and no history of breast cancer were significantly associated with high BPE. Among premenopausal patients, no association was found between BPE and time from last menstrual period to CEM. Overall interreader agreement on BPE was moderate (κ = 0.41; 95% CI, 0.40-0.42). Interreader agreement on tissue density was substantial (κ = 0.67; 95% CI, 0.66-0.69). CONCLUSION. Women with dense breasts, premenopausal status, and younger age are more likely to have greater BPE. Targeting CEM to the last menstrual period is not indicated.

Contrast-enhanced Mammography: Current Applications and Future Directions.

Contrast-enhanced mammography (CEM) is a developing modality used for the workup and management of breast cancer. Although diagnostic imaging modalities such as mammography and US have historically been the mainstays of initial breast cancer workup, recent advances in breast MRI have allowed better disease evaluation. However, MRI is not always readily available, can be time consuming, and is contraindicated in certain patients. CEM is an alternative to US and MRI, and it can be used to obtain contrast material-enhanced information and standard mammograms simultaneously. A CEM examination is shorter than that of MRI, and the modalities have similar rates of sensitivity to detect lesions. CEM also costs less than MRI. The authors evaluate clinical uses of CEM and discuss the literature supporting these indications.©RSNA, 2019.

Comparison of Contrast-Enhanced Mammography With Conventional Digital Mammography in Breast Cancer Screening: A Pilot Study.

PURPOSE: To perform a pilot evaluation of contrast-enhanced mammography (CEM) for screening to determine whether it can improve accuracy and reader confidence in diagnosis. METHODS AND MATERIALS: This institutional review board-approved reader study was comprised of 64 de-identified CEM cases acquired from December 1, 2014, to June 7, 2016, including 48 negative, 5 biopsy-proven benign, and 11 biopsy-proven malignancies. Negative cases were followed for at least 2 years without evidence of cancer. Ten breast imagers of varying experience first rated the low-energy (LE) mammogram and then the CEM examination using BI-RADS categories and a 5-point Likert scale for confidence in diagnosis. RESULTS: There were 635 out a total possible 640 complete reader interpretations included in this analysis. The remaining five incomplete interpretations were excluded. Median sensitivity and specificity improved with the addition of CEM (sensitivity: 0.86 [95% confidence interval {CI}: 0.74-0.95] versus 1 [95% CI: 0.83-1.00], specificity: 0.85 [95% CI: 0.64-0.94] versus 0.88 [95% CI: 0.80-0.92]). Individual receiver operating characteristic curves showed significant improvement with CEM (mean area under the curve increase = 0.056 [95% CI: 0.015-0.097], P = .002). The addition of CEM significantly improved average confidence in 5 of 10 readers when compared with LE (P < .0001) and improved pooled confidence across all tissue density categories, except the almost entirely fatty category. There was a trend toward improved confidence with increasing tissue density with CEM. Degree of background parenchymal enhancement did not affect readers' level of improvement in confidence when interpreting CEM. SUMMARY: CEM improved reader performance and confidence compared with viewing only LE, suggesting a role for CEM in breast cancer screening for which larger trials are warranted.

Lysosome biogenesis in health and disease.

This review focuses on the pathways that regulate lysosome biogenesis and that are implicated in numerous degenerative storage diseases, including lysosomal storage disorders and late-onset neurodegenerative diseases. Lysosomal proteins are synthesized in the endoplasmic reticulum and trafficked to the endolysosomal system through the secretory route. Several receptors have been characterized that execute post-Golgi trafficking of lysosomal proteins. Some of them recognize their cargo proteins based on specific amino acid signatures, others based on a particular glycan modification that is exclusively found on lysosomal proteins. Nearly all receptors serving lysosome biogenesis are under the transcriptional control of transcription factor EB (TFEB), a master regulator of the lysosomal system. TFEB coordinates the expression of lysosomal hydrolases, lysosomal membrane proteins, and autophagy proteins in response to pathways sensing lysosomal stress and the nutritional conditions of the cell among other stimuli. TFEB is primed for activation in lysosomal storage disorders but surprisingly its function is impaired in some late-onset neurodegenerative storage diseases like Alzheimer's and Parkinson's, because of specific detrimental interactions that limit TFEB expression or activation. Thus, disrupted TFEB function presumably plays a role in the pathogenesis of these diseases. Multiple studies in animal models of degenerative storage diseases have shown that exogenous expression of TFEB and pharmacological activation of endogenous TFEB attenuate disease phenotypes. These results highlight TFEB-mediated enhancement of lysosomal biogenesis and function as a candidate strategy to counteract the progression of these diseases. This article is part of the Special Issue "Lysosomal Storage Disorders".

CLN8 is an endoplasmic reticulum cargo receptor that regulates lysosome biogenesis.

Organelle biogenesis requires proper transport of proteins from their site of synthesis to their target subcellular compartment1-3. Lysosomal enzymes are synthesized in the endoplasmic reticulum (ER) and traffic through the Golgi complex before being transferred to the endolysosomal system4-6, but how they are transferred from the ER to the Golgi is unknown. Here, we show that ER-to-Golgi transfer of lysosomal enzymes requires CLN8, an ER-associated membrane protein whose loss of function leads to the lysosomal storage disorder, neuronal ceroid lipofuscinosis 8 (a type of Batten disease)7. ER-to-Golgi trafficking of CLN8 requires interaction with the COPII and COPI machineries via specific export and retrieval signals localized in the cytosolic carboxy terminus of CLN8. CLN8 deficiency leads to depletion of soluble enzymes in the lysosome, thus impairing lysosome biogenesis. Binding to lysosomal enzymes requires the second luminal loop of CLN8 and is abolished by some disease-causing mutations within this region. Our data establish an unanticipated example of an ER receptor serving the biogenesis of an organelle and indicate that impaired transport of lysosomal enzymes underlies Batten disease caused by mutations in CLN8.

Trehalose reduces retinal degeneration, neuroinflammation and storage burden caused by a lysosomal hydrolase deficiency.

The accumulation of undegraded molecular material leads to progressive neurodegeneration in a number of lysosomal storage disorders (LSDs) that are caused by functional deficiencies of lysosomal hydrolases. To determine whether inducing macroautophagy/autophagy via small-molecule therapy would be effective for neuropathic LSDs due to enzyme deficiency, we treated a mouse model of mucopolysaccharidosis IIIB (MPS IIIB), a storage disorder caused by deficiency of the enzyme NAGLU (alpha-N-acetylglucosaminidase [Sanfilippo disease IIIB]), with the autophagy-inducing compound trehalose. Treated naglu-/ - mice lived longer, displayed less hyperactivity and anxiety, retained their vision (and retinal photoreceptors), and showed reduced inflammation in the brain and retina. Treated mice also showed improved clearance of autophagic vacuoles in neuronal and glial cells, accompanied by activation of the TFEB transcriptional network that controls lysosomal biogenesis and autophagic flux. Therefore, small-molecule-induced autophagy enhancement can improve the neurological symptoms associated with a lysosomal enzyme deficiency and could provide a viable therapeutic approach to neuropathic LSDs. ABBREVIATIONS: ANOVA: analysis of variance; Atg7: autophagy related 7; AV: autophagic vacuoles; CD68: cd68 antigen; ERG: electroretinogram; ERT: enzyme replacement therapy; GAPDH: glyceraldehyde-3-phosphate dehydrogenase; GFAP: glial fibrillary acidic protein; GNAT2: guanine nucleotide binding protein, alpha transducing 2; HSCT: hematopoietic stem cell transplantation; INL: inner nuclear layer; LC3: microtubule-associated protein 1 light chain 3 alpha; MPS: mucopolysaccharidoses; NAGLU: alpha-N-acetylglucosaminidase (Sanfilippo disease IIIB); ONL: outer nuclear layer; PBS: phosphate-buffered saline; PRKCA/PKCα: protein kinase C, alpha; S1BF: somatosensory cortex; SQSTM1: sequestosome 1; TEM: transmission electron microscopy; TFEB: transcription factor EB; VMP/VPL: ventral posterior nuclei of the thalamus.

Lysosomes and Brain Health.

One of the fundamental properties of the cell is the capability to digest and remodel its own components according to metabolic and developmental needs. This is accomplished via the autophagy-lysosome system, a pathway of critical importance in the brain, where it contributes to neuronal plasticity and must protect nonreplaceable neurons from the potentially harmful accumulation of cellular waste. The study of lysosomal biogenesis and function in the context of common and rare neurodegenerative diseases has revealed that a dysfunctional autophagy-lysosome system is the shared nexus where multiple, interconnected pathogenic events take place. The characterization of pathways and mechanisms regulating the lysosomal system and autophagic clearance offers unprecedented opportunities for the development of polyvalent therapeutic strategies based on the enhancement of the autophagy-lysosome pathway to maintain cellular homeostasis and achieve neuroprotection.

Corrigendum: mTORC1-independent TFEB activation via Akt inhibition promotes cellular clearance in neurodegenerative storage diseases.

This corrects the article DOI: 10.1038/ncomms14338.

mTORC1-independent TFEB activation via Akt inhibition promotes cellular clearance in neurodegenerative storage diseases.

Neurodegenerative diseases characterized by aberrant accumulation of undigested cellular components represent unmet medical conditions for which the identification of actionable targets is urgently needed. Here we identify a pharmacologically actionable pathway that controls cellular clearance via Akt modulation of transcription factor EB (TFEB), a master regulator of lysosomal pathways. We show that Akt phosphorylates TFEB at Ser467 and represses TFEB nuclear translocation independently of mechanistic target of rapamycin complex 1 (mTORC1), a known TFEB inhibitor. The autophagy enhancer trehalose activates TFEB by diminishing Akt activity. Administration of trehalose to a mouse model of Batten disease, a prototypical neurodegenerative disease presenting with intralysosomal storage, enhances clearance of proteolipid aggregates, reduces neuropathology and prolongs survival of diseased mice. Pharmacological inhibition of Akt promotes cellular clearance in cells from patients with a variety of lysosomal diseases, thus suggesting broad applicability of this approach. These findings open new perspectives for the clinical translation of TFEB-mediated enhancement of cellular clearance in neurodegenerative storage diseases.

NADPH oxidase promotes Parkinsonian phenotypes by impairing autophagic flux in an mTORC1-independent fashion in a cellular model of Parkinson's disease.

Oxidative stress and aberrant accumulation of misfolded proteins in the cytosol are key pathological features associated with Parkinson's disease (PD). NADPH oxidase (Nox2) is upregulated in the pathogenesis of PD; however, the underlying mechanism(s) of Nox2-mediated oxidative stress in PD pathogenesis are still unknown. Using a rotenone-inducible cellular model of PD, we observed that a short exposure to rotenone (0.5 µM) resulted in impaired autophagic flux through activation of a Nox2 dependent Src/PI3K/Akt axis, with a consequent disruption of a Beclin1-VPS34 interaction that was independent of mTORC1 activity. Sustained exposure to rotenone at a higher dose (10 µM) decreased mTORC1 activity; however, autophagic flux was still impaired due to dysregulation of lysosomal activity with subsequent induction of the apoptotic machinery. Cumulatively, our results highlight a complex pathogenic mechanism for PD where short- and long-term oxidative stress alters different signaling pathways, ultimately resulting in anomalous autophagic activity and disease phenotype. Inhibition of Nox2-dependent oxidative stress attenuated the impaired autophagy and cell death, highlighting the importance and therapeutic potential of these pathways for treating patients with PD.

Electrophysiological and Histological Characterization of Rod-Cone Retinal Degeneration and Microglia Activation in a Mouse Model of Mucopolysaccharidosis Type IIIB.

Sanfilippo syndrome Type B or Mucopolysaccharidosis IIIB (MPS IIIB) is a neurodegenerative autosomal recessive lysosomal storage disorder in which patients suffer severe vision loss from associated retinopathy. Here we sought to study the underlying retinal functional and morphological changes associated with MPS IIIB disease progression using the established model of MPS IIIB, the B6.129S6-Naglu(tm1Efn)/J mouse line. Electroretinogram (ERG) was recorded from MPS IIIB and wild-type (WT) mice at the age of 28 and 46 weeks, and retinal tissues were subsequently collected for immunohistochemistry analysis. At the 28th week, rod a- and b-wave amplitudes were significantly diminished in MPS IIIB compared to WT mice. The cone a- and b-waves of MPS IIIB mice were not significantly different from those of the control at the 28th week but were significantly diminished at the 46 th week, when MPS IIIB mice showed a major loss of rods and rod bipolar cells in both central and peripheral regions and a minor loss of cones in the periphery. Activation of microglia and neovascularization were also detected in the MPS IIIB retina. The new findings that cones and rod bipolar cells also undergo degeneration, and that retinal microglia are activated, will inform future development of therapeutic strategies.

Papilloma on core biopsy: excision vs. observation.

BACKGROUND: Intraductal papillomas (IPs) are commonly seen breast lesions with variable clinical presentation. For a palpable lesion and/or evidence of cellular atypia and/or pathologic nipple discharge, excision is warranted to rule out adjacent carcinoma, while for asymptomatic IPs lacking atypia current data for excision vs. observation are controversial. We reviewed outcomes of IPs diagnosed at our institution. METHODS: With IRB approval, we reviewed consecutive patients with IPs seen on core biopsy (CBx) between 2005 and 2013. All patients had an excision, with subspecialty breast pathology review of CBx and excisions. The rate of upgrade to cancer on excision was recorded. Differences between atypia and no-atypia groups were determined by two-tailed t test and Fisher's exact test. RESULTS: We identified 97 patients (age range 31-83 years) with IPs on CBx. Among 52 atypical IPs, DCIS was seen in 11 (upgrade 21 %). In 45 IPs without atypia, 3 cancers were seen (upgrade 6. %): 2 had palpable lesions and were found to have DCIS, and 1 invasive cancer was found in a non-palpable mammographically detected BIRADS 4C lesion, whose Cbx result was discordant. If the 2 palpable lesions are excluded, the upgrade rate for IPs without atypia is 2.2 %. CONCLUSIONS: This series shows a low upgrade rate for IP without atypia seen on CBx in the absence of a palpable mass and radiographic/pathologic discordance, suggesting that a surgical biopsy may not be necessary. Further prospective studies to better estimate the upgrade rate for IPs without atypia may be helpful.

2-Hydroxypropyl-ß-cyclodextrin promotes transcription factor EB-mediated activation of autophagy: implications for therapy.

2-Hydroxypropyl-ß-cyclodextrin (HPßCD) is a Food and Drug Administration-approved excipient used to improve the stability and bioavailability of drugs. Despite its wide use as a drug delivery vehicle and the recent approval of a clinical trial to evaluate its potential for the treatment of a cholesterol storage disorder, the cellular pathways involved in the adaptive response that is activated upon exposure to HPßCD are still poorly defined. Here, we show that cell treatment with HPßCD results in the activation of the transcription factor EB, a master regulator of lysosomal function and autophagy, and in enhancement of the cellular autophagic clearance capacity. HPßCD administration promotes transcription factor EB-mediated clearance of proteolipid aggregates that accumulate due to inefficient activity of the lysosome-autophagy system in cells derived from a patient with a lysosomal storage disorder. Interestingly, HPßCD-mediated activation of autophagy was found not to be associated with activation of apoptotic pathways. This study provides a mechanistic understanding of the cellular response to HPßCD treatment, which will inform the development of safe HPßCD-based therapeutic modalities and may enable engineering HPßCD as a platform technology to reduce the accumulation of lysosomal storage material.

A rapid and sensitive method for measuring N-acetylglucosaminidase activity in cultured cells.

A rapid and sensitive method to quantitatively assess N-acetylglucosaminidase (NAG) activity in cultured cells is highly desirable for both basic research and clinical studies. NAG activity is deficient in cells from patients with Mucopolysaccharidosis type IIIB (MPS IIIB) due to mutations in NAGLU, the gene that encodes NAG. Currently available techniques for measuring NAG activity in patient-derived cell lines include chromogenic and fluorogenic assays and provide a biochemical method for the diagnosis of MPS IIIB. However, standard protocols require large amounts of cells, cell disruption by sonication or freeze-thawing, and normalization to the cellular protein content, resulting in an error-prone procedure that is material- and time-consuming and that produces highly variable results. Here we report a new procedure for measuring NAG activity in cultured cells. This procedure is based on the use of the fluorogenic NAG substrate, 4-Methylumbelliferyl-2-acetamido-2-deoxy-alpha-D-glucopyranoside (MUG), in a one-step cell assay that does not require cell disruption or post-assay normalization and that employs a low number of cells in 96-well plate format. We show that the NAG one-step cell assay greatly discriminates between wild-type and MPS IIIB patient-derived fibroblasts, thus providing a rapid method for the detection of deficiencies in NAG activity. We also show that the assay is sensitive to changes in NAG activity due to increases in NAGLU expression achieved by either overexpressing the transcription factor EB (TFEB), a master regulator of lysosomal function, or by inducing TFEB activation chemically. Because of its small format, rapidity, sensitivity and reproducibility, the NAG one-step cell assay is suitable for multiple procedures, including the high-throughput screening of chemical libraries to identify modulators of NAG expression, folding and activity, and the investigation of candidate molecules and constructs for applications in enzyme replacement therapy, gene therapy, and combination therapies.

Modality-specific axonal regeneration: toward selective regenerative neural interfaces.

Regenerative peripheral nerve interfaces have been proposed as viable alternatives for the natural control of robotic prosthetic devices. However, sensory and motor axons at the neural interface are of mixed sub-modality types, which difficult the specific recording from motor axons and the eliciting of precise sensory modalities through selective stimulation. Here we evaluated the possibility of using type specific neurotrophins to preferentially entice the regeneration of defined axonal populations from transected peripheral nerves into separate compartments. Segregation of mixed sensory fibers from dorsal root ganglion neurons was evaluated in vitro by compartmentalized diffusion delivery of nerve growth factor (NGF) and neurotrophin-3 (NT-3), to preferentially entice the growth of TrkA+ nociceptive and TrkC+ proprioceptive subsets of sensory neurons, respectively. The average axon length in the NGF channel increased 2.5-fold compared to that in saline or NT-3, whereas the number of branches increased threefold in the NT-3 channels. These results were confirmed using a 3D "Y"-shaped in vitro assay showing that the arm containing NGF was able to entice a fivefold increase in axonal length of unbranched fibers. To address if such segregation can be enticed in vivo, a "Y"-shaped tubing was used to allow regeneration of the transected adult rat sciatic nerve into separate compartments filled with either NFG or NT-3. A significant increase in the number of CGRP+ pain fibers were attracted toward the sural nerve, while N-52+ large-diameter axons were observed in the tibial and NT-3 compartments. This study demonstrates the guided enrichment of sensory axons in specific regenerative chambers, and supports the notion that neurotrophic factors can be used to segregate sensory and perhaps motor axons in separate peripheral interfaces.