Catabolism of lysosome-related organelles in color-changing spiders supports intracellular turnover of pigments

Pigment organelles of vertebrates belong to the lysosome-related organelle (LRO) family, of which melanin-producing melanosomes are the prototypes. While their anabolism has been extensively unraveled through the study of melanosomes in skin melanocytes, their catabolism remains poorly known. Here, we tap into the unique ability of crab spiders to reversibly change body coloration to examine the catabolism of their pigment organelles. By combining ultrastructural and metal analyses on high-pressure frozen integuments, we first assess whether pigment organelles of crab spiders belong to the LRO family and second, how their catabolism is intracellularly processed. Using scanning transmission electron microscopy, electron tomography, and nanoscale Synchrotron-based scanning X-ray fluorescence, we show that pigment organelles possess ultrastructural and chemical hallmarks of LROs, including intraluminal vesicles and metal deposits, similar to melanosomes. Monitoring ultrastructural changes during bleaching suggests that the catabolism of pigment organelles involves the degradation and removal of their intraluminal content, possibly through lysosomal mechanisms. In contrast to skin melanosomes, anabolism and catabolism of pigments proceed within the same cell without requiring either cell death or secretion/phagocytosis. Our work hence provides support for the hypothesis that the endolysosomal system is fully functionalized for within-cell turnover of pigments, leading to functional maintenance under adverse conditions and phenotypic plasticity. First formulated for eye melanosomes in the context of human vision, the hypothesis of intracellular turnover of pigments gets unprecedented strong support from pigment organelles of spiders.

How and why animals produce their colors are fundamental questions in biology. In cases where pigments are involved, they are usually synthesized and stored in specialized intracellular organelles (1). Bagnara et al. (2) postulated in 1979 that all pigment organelles of vertebrates derive from a common primordial organelle. Since then, an increasing body of evidence has shown that pigment organelles, from mammal melanosomes to snake pterinosomes, belong to the lysosome-related organelle (LRO) family (3, 4). LROs intersect the endolysosomal system (5), which forms a complex and active network of membrane-bound compartments produced by endocytic and secretory pathways (6). Studying the subcellular aspect of LROs in relation to pigmentation is therefore a critical step to understand animal coloration.Detailed investigations of intracellular processes and trafficking leading to the biogenesis of pigment LROs have been largely performed on mammalian melanosomes (3, 5). They revealed that skin melanocytes divert components of the endolysosomal system to progressively generate melanosome precursors derived from endosomes, bearing intraluminal vesicles (ILVs) and amyloid fibrils, and then mature pigmented melanosomes that are transferred to keratinocytes. Melanosome formation is controlled by a range of genes involved in the endolysosomal system of mammals (3, 5). The finding of homologous genes controlling pterinosomes, iridisomes, and ommochromasomes of snakes and insects, among others, has led to a general model of pigment organelle formation (3, 4, 7). However, coloration involves not only pigmentation phases but also, bleaching phases that lead to pigment removal, a process that is far less understood, even for melanosomes (8).While bleaching can result from death of pigment-containing cells, such as during peeling (9), this process is also compatible with pigment cells remaining alive (10). The latter phenomenon questions how pigment cells accommodate both the production and the removal of pigment organelles, as well as whether recycling pathways connecting the two phases exist. The occurrence of within-cell melanosomal degradation as a common physiological process required for melanin turnover has long been debated (8, 11–13). Here, turnover is defined as a pigmentation–depigmentation–repigmentation cycle at the cellular scale, without requiring the reuse of organellar materials. Evidence for physiological degradations of pigment LROs remains scarce [but see the studies on differences in human skin pigmentation (14, 15)], which we attribute to the rarity of biological systems displaying active and concomitant production and removal of pigments within a single cell and a lack of studies focusing on this phase.Color-changing crab spiders can dynamically match the flower color on which they hunt (16, 17). They do so by reversibly changing their body coloration over the course of a few days, up to weeks, between white and yellow (18). This slow morphological color changes results from the metabolism of yellow pigments, thought to be ommochromes, in integument cells (10, 18, 19). Ommochromes are pigments deriving from tryptophan through the formation of colored kynurenines and transient precursors (7, 20). They are involved in various color-changing species, including dragonflies and cephalopods (21–23). The crab spider is the best understood nonmodel system among the ones displaying morphological color changes in terms of pigment organelle metabolism, ranging from locusts to planarians (24, 25). During yellowing (i.e., anabolic phase), pigments are deposited within specialized intracellular organelles, whose intracellular origin remains undetermined (19). During bleaching, pigment organelles are thought to undergo an autocatalytic process and to recycle their membrane for another cycle of yellowing (10). However, ultrastructural and chemical evidence for this recycling process is scarce, and we do not fully comprehend how both anabolism and catabolism of ommochromes and pigment organelles of crab spiders fit within intracellular trafficking pathways (7). Testing the hypothesis that these pigment organelles are members of the LRO family may help position them into the endolysosomal system, which in return, might provide insights into the roles of the endolysosomal system in both pigmentation and bleaching phases, as well as in reversibility.LROs are defined as being morphologically distinct from lysosomes, containing a subset of cell type–specific contents necessary for their function and being in many instances secretory organelles. However, LROs also share features with lysosomes such as the presence of lysosomal-associated membrane proteins, hydrolases, and an acidic pH (at least temporary), and some can be accessible via the endocytic pathway (3, 5). In practice, the assignment of organelles to the LRO family is based on genetic defects associated with human diseases, such as Hermansky–Pudlak, Griscelli, and Chediak–Higashi syndromes (3–5). However, such genetic dependence cannot be tested in spiders because they are in most cases not genetically tractable. Therefore, other markers of their secretory/endocytic nature should be assessed. LROs often bear ultrastructural signatures of their intracellular origin, such as endosomal/Golgi connections, ILVs, physiological amyloid fibrils, and membrane tubulations (6). Conversely, LROs act as important regulators of metals by storing and releasing them in a dynamic manner (26). Specific techniques are required to retain such labile signatures as membrane tubulations and metals. For relatively thick tissues, this is best done by high-pressure freezing (HPF), which avoids artifacts produced by chemical fixatives (27). As an example, electron microscopy (EM) combined with HPF successfully revealed the morphological details of LROs like melanosomes (28) and Weibel–Palade bodies (29, 30) and their close contacts with other organelles (28). Given the submicrometer size of pigment organelles (19), detection of metals in their lumen can only be tackled by highly sensitive and spatially resolved nanoimaging methods (31), like scanning Synchrotron X-ray fluorescence (SXRF). SXRF is a state-of-the-art chemical imaging technique to map multiple trace elements with subpart per million sensitivity down to a few tens of nanometers resolution (32–34). Therefore, to reveal the intracellular origin of pigment organelles and their catabolism, we exploited the combination of EM and SXRF on white, yellow, and bleaching crab spiders fixed in a near-native state by HPF (SI Appendix discusses the overall fixation quality; SI Appendix, Fig. S1).     

B‐SPID: An object‐relational database architecture to store,retrieve, and manipulate neuroimaging data

We propose a hardware and software architecture to respond to crucial problems in the neuroimaging field: storage, retrieval, and processing of large datasets. The B‐SPID project, here discussed, concerns the processing of neuroimages and attached components stored in an object‐relational multimedia database management system (DBMS). Advanced bioinformation concepts are exploited in this project such as large scale data storage, high level graphical user interfaces and 3D graphical processing and display of data. Our database implementation is based on standard programming components, runs on several UNIX platforms and is written to be evolutive. Queries on this database are designed to obtain and display from neuroimaging data several types of results (pictures, text, or 3D graphical shapes) on heterogeneous systems. Hum. Brain Mapping 7:136–150, 1999. © 1999 Wiley‐Liss, Inc.     

Radiation dose,chemotherapy and risk of osteosarcoma after solid tumours during childhood

Osteosarcoma is the most frequent second primary cancer occurring during the first 20 years following treatment for a solid cancer in childhood. Using a cohort study of children treated for a solid cancer, we investigated the incidence and etiology of osteosarcoma as a second malignant neoplasm after childhood cancer in a cohort and a case-control study. We analysed the relationship between the local dose of radiation and the risk of osteosarcoma, taking into account chemotherapy received. A cohort study of 4,400 3-year survivors of a first solid cancer during childhood diagnosed in France or the United Kingdom, between 1942 and 1986, revealed 32 subsequent osteosarcomas. In a nested case-control study, we matched 32 cases and 160 controls for sex, type of first cancer, age at first cancer and the duration of follow-up. Parameters studied were the incidence of osteosarcoma, the cumulative local dose of irradiation and the cumulative dose of chemotherapy received by cases and controls. The risk of a osteosarcoma was found to be a linear function of the local dose of radiation (excess relative risk per gray = 1.8), and was found to increase with the number of moles of electrophilic agents per square meter but not with other drugs. No interaction was noted between radiotherapy and chemotherapy. Bilateral retinoblastoma, Ewing's sarcoma and soft tissue sarcoma were found to render patients susceptible to a higher risk of developing an osteosarcoma as a second malignant neoplasm. We recommend long-term surveillance of patients who were treated during childhood for bilateral retinoblastoma, Ewing's sarcoma, soft tissue sarcoma, as well as other first cancer treated with radiotherapy plus high doses of chemotherapy, without focusing exclusively on the radiation field. Int. J. Cancer 77:370–377, 1998. © 1998 Wiley-Liss, Inc.     

Does Change over Time in Delusional Beliefs as Measured with PDI Predict Change over Time in Belief Flexibility Measured with MADS?

Psychiatric Quarterly - Delusional beliefs and their behavioral consequences are predominant symptoms in patients with psychosis and play an important role in the treatment. Delusional beliefs are...     

Prevalence and risk factors for adenomas in the ileal pouch and the afferent loop after restorative proctocolectomy for patients with familial adenomatous polyposis

Background

Restorative proctocolectomy with ileal pouch–anal anastomosis (IPAA) has become the treatment of choice in familial adenomatous polyposis (FAP) to prevent the risk of colorectal cancer. However, it currently is recognized that adenomas may develop in the ileal pouch. The risk of adenoma occurring in the afferent ileal loop above the pouch is less clearly identified. This study aimed to evaluate the difference in prevalence of adenomas between the ileal pouch and the afferent ileum after IPAA in FAP.

Methods

The study analyzed 442 endoscopies performed between 2003 and 2008 for 139 FAP patients. The patients had undergone an IPAA in 118 cases, an ileorectal anastomosis in 13 cases, or an ileostomy in 8 cases.

Results

Among the 118 IPAA patients, 57 (48.3 %) had pouch adenomas a median of 15 years after surgery. The risk factors for pouch adenomas were delay since pouch construction [odds ratio (OR), 1.11; p = 0.016] and presence of advanced duodenal adenomas (OR, 4.35; p = 0.011). Seven patients had pouch adenomas with high-grade dysplasia. Only nine patients had afferent ileal loop adenomas (6.5 %). The only significant risk factor for ileal adenomas was the presence of pouch adenomas (OR, 2.16; p = 0.007).

Conclusion

After restorative proctocolectomy in FAP, adenoma recurrence is frequent in the pouch, with a higher risk for patients with advanced duodenal adenomas and an increasing risk over time, whereas adenomas are rarely found in the afferent ileal loop. This finding may help to propose redo ileal pouch anal anastomosis if required.     

Screening for Lynch Syndrome in Colorectal Cancer: Are We Doing Enough?

Purpose

The purpose of this study was to assess the efficacy of screening for the detection of Lynch syndrome (LS) in an unselected population undergoing surgery for a colorectal cancer.

Methods

A total of 1,040 patients were prospectively included between 2005 and 2009. LS screening modalities included the Bethesda criteria, immunochemistry (IHC) for MLH1, MSH2, and MSH6, and microsatellite instability (MSI) by using pentaplex markers. Promoter methylation was assessed in tumors with a loss of MLH1 expression. Gene sequencing was offered to patients with abnormal IHC or MSI status without promoter methylation.

Results

A total of 105 patients had an abnormal result: 102 (9.8%) exhibited a loss of protein on IHC and 98 (9.4%) had MSI. A discordant result was observed in 10 patients with eventual proven LS in 6 patients. Loss of MLH1 (n?=?64) was due to promoter methylation in 43 patients (67.2%). Overall, of 62 patients with an abnormal result, 38 had genetic sequencing leading to 25 (65.8%) identified with a germ-line mutation. Loss of MSH2 on IHC was associated with a mutation in 78.3% (18 of 23) of cases. Among the 62 patients with abnormal results, 23 (37.1%) did not meet the Bethesda criteria.

Conclusions

Strict application of the Bethesda criteria does not lead to identification of all patients with LS. IHC and MSI testing are complementary methods and should be used in association to identify potential LS patients.     

A case of atypical neurological symptoms and high cerebro-spinal fluid acetone level due to transdermal exposure to isopropanol

<正>Dear editor, Isopropanol, also named 2-propanol or isopropyl alcohol(IPA), is a colorless, volatile liquid found in numerous household chemicals, such as cleaners and disinfectants, which typically contain a 70% solution of IPA in water. IPA is also extensively used in industry and laboratories as a solvent.     

Cell-free DNA technologies for the analysis of brain cancer

Survival for glioma patients has shown minimal improvement over the past 20 years. The ability to detect and monitor gliomas relies primarily upon imaging technologies that lack sensitivity and specificity, especially during the post-surgical treatment phase. Treatment-response monitoring with an effective liquid-biopsy paradigm may also provide the most facile clinical scenario for liquid-biopsy integration into brain-tumour care. Conceptually, liquid biopsy is advantageous when compared with both tissue sampling (less invasive) and imaging (more sensitive and specific), but is hampered by technical and biological problems. These problems predominantly relate to low concentrations of tumour-derived DNA in the bloodstream of glioma patients. In this review, we highlight methods by which the neuro-oncological scientific and clinical communities have attempted to circumvent this limitation. The use of novel biological, technological and computational approaches will be explored. The utility of alternate bio-fluids, tumour-guided sequencing, epigenomic and fragmentomic methods may eventually be leveraged to provide the biological and technological means to unlock a wide range of clinical applications for liquid biopsy in glioma.Subject terms: CNS cancer, Next-generation sequencing, Biomarkers