Lipofuscin Deposition Research Articles

An ABCA4 loss-of-function mutation causes a canine form of Stargardt disease.

Autosomal recessive retinal degenerative diseases cause visual impairment and blindness in both humans and dogs. Currently, no standard treatment is available, but pioneering gene therapy-based canine models have been instrumental for clinical trials in humans. To study a novel form of retinal degeneration in Labrador retriever dogs with clinical signs indicating cone and rod degeneration, we used whole-genome sequencing of an affected sib-pair and their unaffected parents. A frameshift insertion in the ATP binding cassette subfamily A member 4 (ABCA4) gene (c.4176insC), leading to a premature stop codon in exon 28 (p.F1393Lfs*1395), was identified. In contrast to unaffected dogs, no full-length ABCA4 protein was detected in the retina of an affected dog. The ABCA4 gene encodes a membrane transporter protein localized in the outer segments of rod and cone photoreceptors. In humans, the ABCA4 gene is associated with Stargardt disease (STGD), an autosomal recessive retinal degeneration leading to central visual impairment. A hallmark of STGD is the accumulation of lipofuscin deposits in the retinal pigment epithelium (RPE). The discovery of a canine homozygous ABCA4 loss-of-function mutation may advance the development of dog as a large animal model for human STGD.

Evaluation of the effects of three sulfa sweeteners on the lifespan and intestinal fat deposition in C. elegans.

High sugar content in beverage or food can affect the aging process, and thus natural/artificial sweeteners are widely used as substitutes. However, whether sweeteners have such adverse effects as sugar remains to be clarified. Therefore, in the current study, three sulfa sweeteners, namely, saccharin sodium salt hydrate (SAC2), sodium cyclamate (CYC3) and acesulfame potassium (AceK4) were evaluated for their effects on the lifespan, deposition of lipofuscin, exercise activity, food intake, and intestinal fat deposition (IFD5) of Caenorhabditis elegans (C. elegans6). It was shown that SAC at 0.3 and 10 mg/mL shortened the lifespan of C. elegans and impaired the exercise capacity, while at other concentrations no significant effects were observed. In contrast, CYC at 0.1, 1 and 10 mg/mL prolonged the lifespan of C. elegans. On the other hand, AceK at 1 mg/mL increased the lifespan of C. elegans, and could decrease both lipofuscin deposition and IFD in a dose-dependent manner. Taken together, these results indicated that although SAC, CYC, and AceK all belong to the sulfa sweeteners, each has distinct effects on different physiological activities associated with aging, at least in C. elegans.

P16Ink4a deletion in cells of the intervertebral disc affects their matrix homeostasis and senescence associated secretory phenotype without altering onset of senescence

Intervertebral disc degeneration is an important contributor to chronic low back and neck pain. Although many environmental and genetic factors are known to contribute to disc degeneration, age is still the most significant risk factor. Recent studies have shown that senescence may play a role in age-related disc degeneration and matrix catabolism in humans and mouse models. Clearance of p16Ink4a-positive senescent cells reduces the degenerative phenotype in many age-associated diseases. Whether p16Ink4a plays a functional role in intervertebral disc degeneration and senescence is unknown. We first characterized the senescence status of discs in young and old mice. Quantitative histology, gene expression and a novel p16tdTom reporter mice showed an increase in p16Ink4a, p21 and IL-6, with a decrease in Ki67 with aging. Accordingly, we studied the spinal-phenotype of 18-month-old mice with conditional deletion of p16Ink4a in the disc driven by Acan-CreERT2 (cKO). The analyses of discs of cKO and age-matched control mice showed little change in cell morphology and tissue architecture. The cKO mice exhibited changes in functional attributes of aggrecan as well as in collagen composition of the intervertebral disc. While cKO discs exhibited a small decrease in TUNEL positive cells, lineage tracing experiments using ZsGreen reporter indicated that the overall changes in cell fate or numbers were minimal. The cKO mice maintained expression of NP-cell phenotypic markers CA3, Krt19 and GLUT-1. Moreover, in cKO discs, levels of p19Arf and RB were higher without alterations in Ki67, γH2AX, CDK4 and Lipofuscin deposition. Interestingly, the cKO discs showed lower levels of SASP markers, IL-1β, IL-6, MCP1 and TGF-β1. These results show that while, p16Ink4a is dispensable for induction and maintenance of senescence, conditional loss of p16Ink4a reduces apoptosis, limits the SASP phenotype and alters matrix homeostasis of disc cells.

Structural differences in the diaphragm of patients following controlled vs assisted and spontaneous mechanical ventilation.

Ventilator-induced diaphragm dysfunction or damage (VIDD) is highly prevalent in patients under mechanical ventilation (MV), but its analysis is limited by the difficulty of obtaining histological samples. In this study we compared diaphragm histological characteristics in Maastricht III (MSIII) and brain-dead (BD) organ donors and in control subjects undergoing thoracic surgery (CTL) after a period of either controlled or spontaneous MV (CMV or SMV). In this prospective study, biopsies were obtained from diaphragm and quadriceps. Demographic variables, comorbidities, severity on admission, treatment, and ventilatory variables were evaluated. Immunohistochemical analysis (fiber size and type percentages) and quantification of abnormal fibers (a surrogate of muscle damage) were performed. Muscle samples were obtained from 35 patients. MSIII (n = 16) had more hours on MV (either CMV or SMV) than BD (n = 14) and also spent more hours and a greater percentage of time with diaphragm stimuli (time in assisted and spontaneous modalities). Cross-sectional area (CSA) was significantly reduced in the diaphragm and quadriceps in both groups in comparison with CTL (n = 5). Quadriceps CSA was significantly decreased in MSIII compared to BD but there were no differences in the diaphragm CSA between the two groups. Those MSIII who spent 100h or more without diaphragm stimuli presented reduced diaphragm CSA without changes in their quadriceps CSA. The proportion of internal nuclei in MSIII diaphragms tended to be higher than in BD diaphragms, and their proportion of lipofuscin deposits tended to be lower, though there were no differences in the quadriceps fiber evaluation. This study provides the first evidence in humans regarding the effects of different modes of MV (controlled, assisted, and spontaneous) on diaphragm myofiber damage, and shows that diaphragm inactivity during mechanical ventilation is associated with the development of VIDD.

SA71ALTERNATIVE SPLICING AND ISOFOROM DIVERSITY IN THE DEVELOPING BRAIN: RELEVANCE FOR NEUROPSYCHIATRIC DISEASE

Progranulin deficiency due to heterozygous null mutations in the GRN gene is a common cause of familial frontotemporal lobar degeneration (FTLD), while homozygous loss-of-function GRN mutations cause neuronal ceroid lipofuscinosis (NCL). Aged progranulin-knockout mice display highly exaggerated lipofuscinosis, microgliosis, and astrogliosis, as well as mild cell loss in specific brain regions. Progranulin is a secreted glycoprotein expressed in both neurons and microglia, but not astrocytes, in the brain. We generated conditional progranulin-knockout mice that lack progranulin in nestin-expressing cells (Nes-cKO mice), which include most neurons as well as astrocytes. We confirmed near complete knockout of progranulin in neurons in Nes-cKO mice, while microglial progranulin levels remained similar to that of wild-type animals. Overall brain progranulin levels were reduced by about 50% in Nes-cKO, and no Grn was detected in primary Nes-cKO neurons. Nes-cKO mice aged to 12 months did not display any increase in lipofuscin deposition, microgliosis, or astrogliosis in the four brain regions examined, though increases were observed for most of these measures in Grn-null animals. We conclude that neuron-specific loss of progranulin is not sufficient to cause similar neuropathological changes to those seen in constitutive Grn-null animals. Our results suggest that increased lipofuscinosis and gliosis in Grn-null animals are not caused by intrinsic progranulin deficiency in neurons, and that microglia-derived progranulin may be sufficient to maintain neuronal health and homeostasis in the brain.

Novel mutations in CLN6 cause late-infantile neuronal ceroid lipofuscinosis without visual impairment in two unrelated patients

CLN6 is a transmembrane protein located in the endoplasmic reticulum that is involved in lysosomal acidification. Mutations in CLN6 cause late-infantile neuronal ceroid lipofuscinosis (LINCL), and teenage and adult onset NCL without visual impairment. Here we describe two pediatric patients with LINCL from unrelated families who were evaluated at the National Institutes of Health. Both children exhibited typical phenotypes associated with LINCL except that they lacked the expected visual impairment. Whole exome sequencing identified novel biallelic mutations in CLN6, i.e., c.218-220dupGGT (p.Trp73dup) and c.296A > G (p.Lys99Arg) in Proband 1 and homozygous c.723G > T (p.Met241Ile) in Proband 2. Expression analysis in dermal fibroblasts showed a small increase in CLN6 protein levels. Electron micrographs of these fibroblasts demonstrated large numbers of small membrane-bound vesicles, in addition to lipofuscin deposits. LysoTracker™ Red intensity was increased in fibroblasts from both patients. This study supports a role for CLN6 in lysosomal homeostasis, and highlights the importance of considering CLN6 mutations in the diagnosis of Batten Disease even in patients with normal vision.

Brown Bowel Syndrome Presenting as a Small Bowel Obstruction

Abstract Brown bowel syndrome (BBS) is a rare condition characterized by lipofuscin deposition in the muscularis propria of the bowel. It is associated with vitamin E deficiency and malabsorptive syndromes. We report a case of BBS in a patient with previously undiagnosed primary intestinal lymphangiectasia. The patient had a longstanding history of malnutrition as well as bilateral lower extremity lymphedema and chylous ascites. He presented to our hospital with small bowel obstruction (SBO). Initial magnetic resonance enterography revealed a dilated duodenum, which when biopsied revealed only lymphangiectasia and lymphedema. He was managed conservatively and discharged following resolution of the obstruction. The patient presented again 1 month later with SBO that failed to resolve with conservative treatment. Subsequent exploratory laparotomy discovered a portion of pale, edematous, aperistaltic jejunum. The specimen was resected, and pathologic examination revealed brown granular pigment deposition within the muscularis propria, which was highlighted by periodic acid–Schiff and lipofuscin stains, diagnostic of BBS. Nutritional workup revealed multiple deficiencies. The patient's clinical findings and history were suggestive of primary intestinal lymphangiectasia with resulting nutritional deficiencies leading to BBS. Outpatient follow-up after recovery revealed an elevated 24-hour fecal α1 antitrypsin, which supports this diagnosis. The patient has since improved following adherence to a low-fat diet. The combined clinical, radiographic, and pathologic findings in this case allowed for the resolution of this complex diagnostic dilemma. Although rare, BBS remains an important consideration in the differential diagnosis of a patient with SBO and history of chylous ascites and should be viewed as a symptom of underlying treatable nutritional deficiency.

Transient acceleration of autophagic degradation by pharmacological Nrf2 activation is important for retinal pigment epithelium cell survival

Non-exudative age-related macular degeneration (AMD) is mainly caused by the accumulation of lipofuscin and drusen on the retinal pigment epithelium (RPE). Both oxidative stress and autophagic dysfunction accelerate the deposition of lipofuscin at the RPE. One of the key regulators in the response against oxidative stress is the NF-E2-Related Factor 2 (Nrf2)-kelch like ECH associated protein 1 (Keap1) axis, which is also closely associated with the autophagy pathway. Nrf2 activation upregulates the expression levels of certain anti-oxidative enzymes [e.g. Heme oxygenase-1 (HO-1)], which attenuates oxidative damage. However, until now, the relationship between cytoprotective effects of Nrf2 activation and autophagic degradation remain unclear. To address these questions, we investigated the effects of a novel Nrf2 activator, RS9, on RPE damage. We found that RS9 protected ARPE-19 cells against NaIO3-induced oxidative damage, and that the protective effects of RS9 were inhibited by co-treatment with zinc protoporphyrin, an HO-1 inhibitor. Next, we examined the involvement of autophagic degradation in the protective effects of RS9. Co-treatment with RS9 and chloroquine, a lysosomal acidification inhibitor, inhibited the protective effect. Furthermore, western blotting and immunostaining showed that RS9 accelerated autophagy flux and induced transient upregulation of p62 [also known as sequestosome 1 (SQSTM1)]. Co-treatment with chloroquine and RS9 also inhibited the degradation of autophagosomes. Transient upregulation of SQSTM1 by RS9 was unaltered by HO-1 knockdown using siRNA. RS9 and chloroquine had the same actions in light damaged adult zebrafish retina as those in vitro. In conclusion, we clarified the relationship between acceleration of the autophagy pathway and the cytoprotective effects of Nrf2 activation in RPE cells and zebrafish retina. These findings indicated that Nrf2 activation could be a promising therapeutic approach for non-exudative AMD by supporting RPE maintenance.

Atypical presentation of Best Disease

Abstract Clinical case A 43-year-old man was treated for reduced visual acuity, initially attributed to strabismic amblyopia. On fundus examination, bilateral neurosensory detachments (NSD) were observed in posterior pole, surrounded by deposits of lipofuscin. His 3-year-old son was also examined and circumscribed NSD was observed with the presence of pseudohypopyon in OD and a fibrosis scar in OS. The Arden ratio was decreased in electrooculography (EOG) in both patients, and genetic studies revealed a single mutation of the BEST1 gene. Discussion The existence of extensive bilateral NSD may be an unusual form of presentation of Best disease. Family history, EOG, and genetic study supported this diagnosis.

A Sequential Study of Age-Related Lipofuscin Accumulation in Hippocampus and Striate Cortex of Rats

Background: The age-pigment, lipofuscin that accumulates in cells intrinsically and progressively with age is considered as the hallmark of aging. This accumulation is more prominent in post-mitotic cells like neurons and also appears in glia. Purpose: The aim of the present study was to assess the age-associated occurrence and distribution pattern of lipofuscin both in neurons and microglia in various regions of hippocampus and striate cortex. Methods: The accumulation pattern of lipofuscin in hippocampus and striate cortex was observed in the female Wistar rats of 6 age groups, that is, 3, 6, 12, 18, 24, and 30 months using the autofluorescent property of lipofuscin and its specific localization in neurons and microglia by dual immunolabeling with NeuN and Iba-1 antibodies respectively. Cytoarchitectural and the morphological age-related changes were observed with cresyl violet staining. Results: Lipofuscin pigments accumulate progressively through the normal aging process in hippocampus and striate cortex. However, in hippocampus, lipofuscin accumulates in a region-specific manner with the highest accumulation observed in cornu ammonis (CA) 1 and 3 subregions. Furthermore, the lipofuscin accumulation was also observed in microglia in the senile brains both in the hippocampus and striate cortex. Conclusion: The progressive deposition of lipofuscin could result in cellular dysfunction. This encouraged us to forward the idea that microglia not only participate in the removal of the pigment from the neurons but also accumulate these waste products in itself. Such senescent microglia may contribute to age- related neurodegeneration as they lose the neuroprotective potential due to oxidative stress and thus may be unable to effectively phagocytose the age pigment.

Морфологические доказательства ишемической природы фиброза предстательной железы при классическом синдроме хронической тазовой боли/хроническом простатите IIIБ

To examine the structure of the prostate tissue in patients with III B chronic prostatitis (CP) and chronic pelvic pain syndrome (CPPS). The study analyzed transrectal fine-needle biopsy specimens of 10 patients with the verified diagnosis of chronic pelvic pain syndrome/category III B chronic prostatitis (CPPS/IIIB CP) according to the National Institutes of Health classification. Tissues were examined using light and electron microscopy, and immunohistochemical study of the expression of CD31, CD34, NSE and S-100 markers. All biopsy specimens of all patients showed fibroplastic changes of the prostate tissue most markedly pronounced in the stroma and muscle fibers in the form of total fibrosis, myofibril atrophy, and extracellular lipofuscin deposition. The examination revealed a significant reduction in the density of microcirculatory bed vessels and arteriolar luminal stenosis, a reduction in the number of nerve fibers, and compression of their fibrous tissue. No inflammatory changes were found in the prostate. In patients with CPPS/IIIB CP, the changes in the prostate at the microscopic and ultrastructural levels are characteristic of severe chronic tissue hypoxia, which leads to the development of fibrosis resulting in stenosis of microcirculatory bed vessels and degenerative changes in nerve fibers and cells. No signs of an inflammatory reaction in the examined tissue were established. Changes in the prostate tissue in CPPS/IIIB CP suggest the presence of chronic pelvic ischemia and exclude its association with inflammation as the main pathological process.

Presentación atípica de la enfermedad de Best

Clinical caseA 43-year-old man was treated for reduced visual acuity, initially attributed to strabismic amblyopia. On fundus examination, bilateral neurosensory detachments (NSD) were observed in posterior pole, surrounded by deposits of lipofuscin. His 3-year-old son was also examined and circumscribed NSD was observed with the presence of pseudohypopyon in OD and a fibrosis scar in OS. The Arden ratio were decreased in electrooculography (EOG) in both patients, and genetic studies revealed a single mutation of the BEST1 gene. DiscussionThe existence of extensive bilateral NSD may be an unusual form of presentation of Best disease. Family history, EOG, and genetic study supported this diagnosis.

Salidroside mediated stabilization of Bcl -xL prevents mitophagy in CA3 hippocampal neurons during hypoxia

Chronic hypoxic stress results in deposition of lipofuscin granules in the CA3 region of hippocampal neurons which contributes to neurodegeneration and accelerated neuronal aging. Oxidative stress and mitophagy during hypoxia are crucial to cause aggregation of these lipofuscin granules in hypoxic neurons. Salidroside, a glucoside derivative of β-Tyrosol, has been reported to protect hypoxic neurons through maintenance of mitochondrial activity. The present study is aimed at investigating the potential of Salidroside in preventing mitophagy during chronic hypoxia and identification of the molecular targets and underlying signaling mechanisms. In-silico analysis for interaction of salidroside with Bcl-xL was carried out using VLife MDS software. The prophylactic efficacy of Salidroside for amelioration of global hypoxia induced neuronal aging was studied in adult male Sprague-Dawley rats exposed to hypobaric hypoxia simulating an altitude of 7600 m for 21 days. Salidroside was supplemented at a daily dose of 25 mg kg−1b.w. p.o. during hypoxic exposure. Ultra-structural and immune-histological studies were conducted to study lipofuscin aggregation and mitophagy. In-silico findings on salidroside mediated stabilization of Bcl-xL were validated by investigating its effect on downstream signaling molecules involved in mitophagy. Administration of Salidroside reduced deposition of lipofuscin in hypoxic CA3 hippocampal neurons and prevented mitophagy. Salidroside stabilizes Bcl-xL in hypoxic neurons resulting in inhibition of PGAM5 phosphatase activity and maintenance of FUNDC1 in phosphorylated state. Salidroside mediated inhibition of pFUNDC1 dephosphorylation prevents FUNDC1-LC3 II interaction which is crucial for mitophagy. The present study demonstrates potential of Salidroside in preventing lipofuscin deposition during chronic hypoxic stress.

Effects of Aging in the Striatum and Substantia Nigra of a Parkinson's Disease Animal Model.

Aging is a multifactorial process associated with functional deficits, and the brain is more prone to developing chronic degenerative diseases such as Parkinson's disease. Several groups have tried to correlate the age-related ultrastructural alterations to the neurodegeneration process using in vivo pharmacological models, but due to the limitations of the animal models, particularly in aged animals, the results are difficult to interpret. In this work, we investigated neurodegeneration induced by rotenone, as a pharmacological model of Parkinson's disease, in both young and aged Wistar rats. We assessed animal mobility, tyrosine hydroxylase staining in the substantia nigra pars compacta (SNpc), and TdT-mediated dUTP-biotin nick end labeling-positive nuclei and reactive oxygen species production in the striatum. Interestingly, the mobility impairment, dopaminergic neuron loss, and elevated number of apoptotic nuclei in the striatum of aged control rats were similar to young rotenone-treated animals. Moreover, we observed many ultrastructural alterations, such as swollen mitochondria in the striatum, and massive lipofuscin deposits in the SNpc of the aged rotenone-treated animals. We conclude that the rotenone model can be employed to explore age-related alterations in the ontogeny that can increase vulnerability in the striatum and SNpc, which may contribute to Parkinson's disease pathogenesis.

Pathological and Clinical Pathological Changes Induced by Four-week, Repeated-dose, Oral Administration of the Wood Preservative Chromated Copper Arsenate in Wistar Rats.

Chromated copper arsenate (CCA) is used as a wood preservative worldwide. Exposure to it may adversely affect human health. Some events have increased human exposure to CCA, including the Great East Japan Earthquake, which generated a large amount of lumber debris from CCA-treated woods. We elucidated the toxicity due to daily exposure to CCA over a 4-week period at doses of 0, 8, 40, and 80 mg/kg/day in Wistar Hannover rats. Chromium (Cr) and arsenic (As), but not copper, were detected in the plasma samples of rats treated with various doses of CCA. Males and females showed sedation, and males had poor body weight gain. The clinical pathologies observed in both sexes included hypochromic and microcytic anemia, hepatic and renal dysfunction, and changes in lipid and glucose levels. Histopathologically, males and females showed forestomach hyperkeratosis, mucosal epithelial hyperplasia in the small intestine, rectal goblet cell hypertrophy, and lipofuscin deposition in the proximal renal tubule. Females showed diffuse hepatocellular hypertrophy with increased 8-hydroxydeoxyguanosine levels. These results indicated that oral administration of CCA mainly affected hematopoietic, gastrointestinal, hepatic, and renal systems owing to the toxic effects of As and/or Cr. Major toxic effects were observed in both sexes receiving 40 and 80 mg/kg/day.

Label-free Imaging of Myocardial Remodeling in Atrial Fibrillation Using Nonlinear Optical Microscopy: A Feasibility Study.

Atrial fibrillation, characterized by rapid disorganized electrical activation of myocardium, is caused by and accompanied by remodeling of myocardial tissue. We applied nonlinear optical microscopy (NLOM) to visualize typical myocardial features and atrial fibrillation effects in order to test anon-destructive imaging technology that in principle can be applied in vivo.Coherent anti-Stokes Raman scattering, endogenous two-photon excited fluorescence, and second harmonic generation were used to inspect unstained human atrial myocardium from three patients who underwent surgical Cox-MAZE procedure with amputation of left atrial appendage. Using NLOM techniques, we collected detailrich pictures of unstained tissue that enable comprehensive characterization of myocardial characteristics like myocyte structure, collagen and lipofuscin deposition, intercalating disc width, and fatty degradation. Development of in vivo application of the NLOM technique may represent a revolutionary approach in characterizing atrial fibrillation associated myocardial remodeling with important implications for therapy individualization and monitoring.

Selective depletion of microglial progranulin in mice is not sufficient to cause neuronal ceroid lipofuscinosis or neuroinflammation

BackgroundProgranulin deficiency due to heterozygous null mutations in the GRN gene are a common cause of familial frontotemporal lobar degeneration (FTLD), while homozygous loss-of-function GRN mutations are thought to be a rare cause of neuronal ceroid lipofuscinosis (NCL). Aged progranulin-knockout (Grn-null) mice display highly exaggerated lipofuscinosis, microgliosis, and astrogliosis, as well as mild cell loss in specific brain regions. In the brain, progranulin is predominantly expressed in neurons and microglia, and previously, we demonstrated that neuronal-specific depletion of progranulin does not recapitulate the neuropathological phenotype of Grn-null mice. In this study, we evaluated whether selective depletion of progranulin expression in myeloid-lineage cells, including microglia, causes NCL-like neuropathology or neuroinflammation in mice.MethodsWe generated mice with progranulin depleted in myeloid-lineage cells by crossing mice homozygous for a floxed progranulin allele to mice expressing Cre recombinase under control of the LyzM promotor (Lyz-cKO).ResultsProgranulin expression was reduced by approximately 50–70% in isolated microglia compared to WT levels. Lyz-cKO mice aged to 12 months did not display any increase in lipofuscin deposition, microgliosis, or astrogliosis in the four brain regions examined, though increases were observed for many of these measures in Grn-null animals. To evaluate the functional effect of reduced progranulin expression in isolated microglia, primary cultures were stimulated with controlled standard endotoxin and cytokine release was measured. While Grn-null microglia display a hyper-inflammatory phenotype, Lyz-cKO and WT microglia secreted similar levels of inflammatory cytokines.ConclusionWe conclude that progranulin expression from either microglia or neurons is sufficient to prevent the development of NCL-like neuropathology in mice. Furthermore, microglia that are deficient for progranulin expression but isolated from a progranulin-rich environment have a normal inflammatory profile. Our results suggest that progranulin acts, at least partly, in a non-cell autonomous manner in the brain.

Localization and functional characterization of the p.Asn965Ser (N965S) ABCA4 variant in mice reveal pathogenic mechanisms underlying Stargardt macular degeneration.

ABCA4 is a member of the superfamily of ATP-binding cassette (ABC) proteins that transports N-retinylidene-phosphatidylethanolamine (N-Ret-PE) across outer segment disc membranes thereby facilitating the removal of potentially toxic retinoid compounds from photoreceptor cells. Mutations in the gene encoding ABCA4 are responsible for Stargardt disease (STGD1), an autosomal recessive retinal degenerative disease that causes severe vision loss. To define the molecular basis for STGD1 associated with the p.Asn965Ser (N965S) mutation in the Walker A motif of nucleotide binding domain 1 (NBD1), we generated a p.Asn965Ser knockin mouse and compared the subcellular localization and molecular properties of the disease variant with wild-type (WT) ABCA4. Here, we show that the p.Asn965Ser ABCA4 variant expresses at half the level of WT ABCA4, partially mislocalizes to the endoplasmic reticulum (ER) of photoreceptors, is devoid of N-Ret-PE activated ATPase activity, and causes an increase in autofluorescence and the bisretinoid A2E associated with lipofuscin deposits in retinal pigment epithelial cells as found in Stargardt patients and Abca4 knockout mice. We also show for the first time that a significant fraction of WT ABCA4 is retained in the inner segment of photoreceptors. On the basis of these studies we conclude that loss in substrate-dependent ATPase activity and protein misfolding are mechanisms underlying STGD1 associated with the p.Asn965Ser mutation in ABCA4. Functional and molecular modeling studies further suggest that similar pathogenic mechanisms are responsible for Tangiers disease associated with the p.Asn935Ser (N935S) mutation in the NBD1 Walker A motif of ABCA1.

Pseudomelanosis Intestini

Introduction: Gastrointestinal melanosis is the term used to describe the accumulation of pigment deposits in the intestinal mucosa. Melanosis coli is a well-known condition associated with the use of anthraceneline laxatives, however melanosis of the small intestine is extremely rare. We report a case of pseudomelanosis inestini involving the pylorus, duodenum, and proximal jejunum in a patient presented with GI bleeding. The clinical significance of this condition is unknown, however gastroenterologists should be aware of its existence. Case Report: A 70 y/o female with a past medical history of atrial fibrillation on warfarin therapy, chronic obstructive pulmonary disease, hypertension, congestive heart failure, and chronic kidney disease stage IIIb, presented with recurrent melena. Her initial EGD revealed a non-bleeding gastric ulcer with a clean ulcer base (Forrest Class III) on the lesser curvature of the gastric antrum, and diffuse duodenal melanosis. She had another episode of melena for which repeated EGD showed same non-bleeding ulcer, and melanosis of the gastric antrum and duodenum. She was treated conservatively and was discharged in stable condition. She presented three weeks later with same complain. Double balloon enteroscopy was performed and revealed melanosis of the duodenum and jejunum (280cm distal to the pylorus). The entire colon and examined portion of the ileum were normal with no evidence of melanosis. Histopathology revealed macrophages containing dark pigmented granules within the lamina propria, which were non-reactive to Perl's iron stain. Discussion: Pseudomelanosis intestine is a benign condition refers to the rare endoscopic appearance of dark pigment deposits in the intestinal mucosa. It can be due to the deposition of melanin-like substances, hemosiderin, lipomelanin, lipofuscin or ferrous sulfate. It usually occurs in patients in their sixth and seventh decade of life with a female predominance. It has been mainly described in patients with certain chronic illnesses, such as chronic kidney disease, hypertension, and GI bleeding. Various medications, such as ferrous sulfate, hydralazine, and furosemide, has been associated with this condition as well. However, the etiologic mechanism, clinical significance, and prognostic and long-term impact of this condition remains unknown. Also, there's no established recommendation regarding the need for therapeutic interventions or regular upper endoscopy survey.Figure: Endoscopic image reveals dark pigment deposits in the pylorus.Figure: Endoscopic image reveals dark pigment deposits in the duodenum.

Severe Melanosis Coli From Rhubarb Supplement

Melanosis coli is a dark pigmentation of colonic wall that is often identified at the time of colonoscopy. We present a second reported case of melanosis coli in a patient taking rhubarb containing herbal laxative. 67-year-old Caucasian female with ulcerative proctosigmoiditis (in clinical remission), sclerosing pancreatitis, and slow transit constipation, noted to have black coloration of the whole colon during a screening colonoscopy. The pigmentation was very dense, limiting the mucosal examination on both standard and peak illumination (Figure 1, panel A/B). This was new from a colonoscopy 2 years prior. Minimal coble stoning was noted in the sigmoid colon. Mucosal biopsies confirmed diagnosis of melanosis coli and chronic colitis with focal mild activity in the sigmoid colon. Her medication list included alprazolam, ascorbic acid, dexlansoprazole, dicyclomine, omega-3 fatty acids, pancrelipase, tramadol and multivitamin and an herbal fiber supplement containing rhubarb. She stopped using rhubarb extract and repeat colonoscopy 2 years later showed complete resolution of melanosis coli (Figure 1, panel C). The cobblestoning of sigmoid colon was also resolved and no active colitis. Melanosis coli is pigmentation of colonic mucosa secondary to lipofuscin deposition in macrophages in the lamina propria. It is considered a result of increased apoptosis. Lysosomal digestion on cellular organelles forms lipofuscin granules. These granules are then taken up by the macrophages. Our patient reported taking rhubarb containing herbal laxative. Rhubarb has anthraquinones which passes unabsorbed through the small bowel and get activated in the colon, where they are converted to active forms. These in turn cause epithelial cell damage leading to altered absorption, secretion and motility. The epithelial cells damage leads to apoptosis. Melanosis coli is considered benign and is reversible on discontinuing implicating agent. Disappearance of the pigment usually occurs within a year after a patient stops taking anthraquinone but microscopic feature could take longer to clear.Figure: A/B - initial colonoscopy showing severe melanosis coli. Fig 1C - Repeat colonoscopy showing improved melanosis coli.