Lysosomal proteolysis and autophagy require presenilin 1 and are disrupted by Alzheimer-related PS1 mutations.

Macroautophagy is a lysosomal degradative pathway essential for neuron survival. Here, we show that macroautophagy requires the Alzheimer's disease (AD)-related protein presenilin-1 (PS1). In PS1 null blastocysts, neurons from mice hypomorphic for PS1 or conditionally depleted of PS1, substrate proteolysis and autophagosome clearance during macroautophagy are prevented as a result of a selective impairment of autolysosome acidification and cathepsin activation. These deficits are caused by failed PS1-dependent targeting of the v-ATPase V0a1 subunit to lysosomes. N-glycosylation of the V0a1 subunit, essential for its efficient ER-to-lysosome delivery, requires the selective binding of PS1 holoprotein to the unglycosylated subunit and the Sec61alpha/oligosaccharyltransferase complex. PS1 mutations causing early-onset AD produce a similar lysosomal/autophagy phenotype in fibroblasts from AD patients. PS1 is therefore essential for v-ATPase targeting to lysosomes, lysosome acidification, and proteolysis during autophagy. Defective lysosomal proteolysis represents a basis for pathogenic protein accumulations and neuronal cell death in AD and suggests previously unidentified therapeutic targets.

TENS-like Stimulation Downregulates Inflammatory Cytokines in a PC-12 Cell Line.

OBJECTIVES: The purpose of this study was to evaluate the effects of transcutaneous electrical nerve stimulation (TENS)-like stimulation on the expression of the proinflammatory cytokines tumor necrosis factor α (TNF-α), interleukin 1ß (IL-1ß), and IL-6 in PC-12 cells, which are commonly used as neuronal cell models. METHODS: Nerve growth factor-differentiated PC-12 cells were exposed to electrical stimulation for 15 minutes at 1 mA, 200 µs, and 100 Hz. Cell lysate from stimulated and control cells was assayed for TNF-α, IL-1ß, and IL-6. In 6 trials, cells were preincubated with the L-type ion channel blocker nicardipine. Cultured cells were also incubated with Alexa Fluor 488 and visualized by fluorescence microscopy to determine the nuclear vs cytoplasmic distribution of the p65 sub-unit of NF-κB RESULTS: Compared with control (unstimulated) cells, the stimulated cells had a downregulation of the assayed cytokines. However, preincubation with the L-type ion channel blocker nicardipine blocked this effect of stimulation. Additionally, it was noted that TENS-like stimulation promoted a relative sequestration of the p65 subunit of NF-κB in the cytoplasm vs the nucleus. CONCLUSIONS: It appears that in this cell line and with these stimulation parameters, TENS-like stimulation attenuated the expression of the assayed proinflammatory cytokines, in part by promoting the relative sequestration of the p65 subunit of NF-κB in the cytoplasm, and that voltage-dependent calcium channels have a role in the cascade of events initiated by the TENS-like stimulation.

Osseointegration of Ti-6Al-4V alloy implants with a titanium nitride coating produced by a PIRAC nitriding technique: a long-term time course study in the rat.

This study examined bone tissue responses to Ti-6Al-4V alloy implants with a hard TiN coating applied by an original powder immersion reaction-assisted coating (PIRAC) nitriding method. Progression of implant fixation in the distal epiphysis and within the medullary cavity of the rat femur was evaluated between 3 days and 6 months postimplantation by scanning electron microscopy, oxytetracycline incorporation, and histochemistry. After 6 months, successful osseointegration was achieved in both epiphyseal and diaphyseal sites. Throughout, implant portions located within the epiphysis remained in close contact with bone trabeculae that gradually engulfed the implant forming a bone collar continuous with the trabecular network of the epiphysis. In the diaphysis, woven bone was first formed within the marrow cavity around the implant and later was replaced by a shell of compact bone around the implant. In general, higher osseointegration rates were measured for TiN-coated versus the uncoated implants, both in the epiphysis and in the diaphysis. In conclusion, our findings indicate an excellent long-term biocompatibility of TiN coatings applied by the PIRAC nitriding technique and superior osteoinductive ability in comparison with uncoated Ti-6Al-4V alloy. Such coatings can, therefore, be considered for improving the corrosion and wear resistance of titanium-based orthopedic implants.

The prevalence and characteristics of HIV/AIDS patients presenting at a chiropractic outpatient clinic in Toronto, Ontario. A retrospective, observational study.

OBJECTIVE: To determine the prevalence and presenting complaints of HIV/AIDS patients attending a chiropractic outpatient teaching clinic in downtown Toronto, and explore their self-reported comorbidities, medications used, and consumption of other complementary health care. METHODS: A random sample was drawn from the entire clinic file collection spanning the years 2007 to 2013. Files were anonymized and coded to ensure confidentiality. RESULTS: A total of 264 files were radomly pulled from approxinately 3750 clinic files. The prevalence of HIV positive patients was 5.7% (15/264), predominantly males, with 3 patients having developed AIDS. Co-infection with Hepatitis B and/or C was identified in 5/15 patients. The most common presenting complaint was neck pain (80%), followed by low back pain (47%) compared to 20% and 43% respectively for the general cohort. Eleven of 15 patients were on antiretroviral treatment (ART); The frequency of comorbidities was 8/15 (53%) however, none were identified as being dominant. In addition to chiropractic, 7/15 patients reported receiving other complementary therapies. CONCLUSIONS: A relatively small proportion of HIV/ AIDS patients were found to be receiving treatments in this downtown chiropractic clinic situated within a community health clinic setting. The principal presenting complaint was neck pain.

OBJECTIF: Établir la prévalence des symptômes des patients séropositifs ou atteints du sida fréquentant un clinique chiropratique d'enseignement au centre-ville de Toronto et étudier les comorbidités autodéclarées, les médicaments utilisés et les soins de santé complémentaires. MÉTHODOLOGIE: On a choisi au hasard des dossiers de patients parmi tous les dossiers de la clinique, à partir de 2007 jusqu'en 2013. Les dossiers ont été anonymisés et codés pour assurer la confidentialité. RÉSULTATS: Au total, 264 dossiers ont été choisis par hasard parmi les quelque 3 750 de la clinique. La prévalence des patients séropositifs était de 5,7 % (15/264); la plupart étaient des hommes, 3 patients avaient développé le sida. Une co-infection par l'hépatite B et (ou) l'hépatite C avait été diagnostiquée chez 5 patients sur 15. Les symptômes les plus fréquents étaient la cervicalgie (80 %) suivie de la lombalgie (47 %); la fréquence de ces symptômes étaient de 20 % et de 43 % respectivement dans la cohorte générale. Onze des 15 patients suivaient un traitement antirétroviral (ARV). La fréquence des comorbidités était de 8 patients sur 15 (53 %), mais aucune n'était considérée comme dominante. Sept patients sur 15 ont déclaré suivre des traitements complémentaires en plus des traitements chiropratiques. CONCLUSIONS: Une proportion relativement petite de patients séropositifs ou atteints du sida recevait des traitements dans cette clinique chiropratique du centre-ville située dans un établissement de soins de santé communautaire. La cervicalgie était le principal symptôme dont se plaignaient les patients.

Chaperones rescue the energetic landscape of mutant CFTR at single molecule and in cell.

Molecular chaperones are pivotal in folding and degradation of the cellular proteome but their impact on the conformational dynamics of near-native membrane proteins with disease relevance remains unknown. Here we report the effect of chaperone activity on the functional conformation of the temperature-sensitive mutant cystic fibrosis channel (∆F508-CFTR) at the plasma membrane and after reconstitution into phospholipid bilayer. Thermally induced unfolding at 37 °C and concomitant functional inactivation of ∆F508-CFTR are partially suppressed by constitutive activity of Hsc70 and Hsp90 chaperone/co-chaperone at the plasma membrane and post-endoplasmic reticulum compartments in vivo, and at single-molecule level in vitro, indicated by kinetic and thermodynamic remodeling of the mutant gating energetics toward its wild-type counterpart. Thus, molecular chaperones can contribute to functional maintenance of ∆F508-CFTR by reshaping the conformational energetics of its final fold, a mechanism with implication in the regulation of metastable ABC transporters and other plasma membrane proteins activity in health and diseases.The F508 deletion (F508del) in the cystic fibrosis transmembrane conductance regulator (CFTR) is the most common CF causing mutation. Here the authors show that cytosolic chaperones shift the F508del channel conformation to the native fold by kinetic and thermodynamic remodelling of the gating energetics towards that of wild-type CTFR.

Protein degradation and aging.

Continuous turnover of intracellular proteins is essential for the maintenance of cellular homeostasis and for the regulation of multiple cellular functions. The first reports showing a decrease in total rates of protein degradation with age are dated more than 50 years ago, when the major players in protein degradation where still to be discovered. The current advances in the molecular characterization of the two main intracellular proteolytic systems, the lysosomal and the ubiquitin proteasome system, offer now the possibility of a systematic search for the defect(s) that lead to the declined activity of these systems in old organisms. We discuss here, in light of the current findings, how malfunctioning of these two proteolytic systems can contribute to different aspects of the phenotype of aging and to the pathogenesis of some age-related diseases.

Missed appendicitis diagnosis: A case report.

OBJECTIVE: The purpose of this case report is to highlight and emphasize the need for an appropriate and thorough list of differential diagnoses when managing patients, as it is insufficient to assume cases are mechanical, until proven non-mechanical. There are over 250,000 cases of appendicitis annually in the United States. Of these cases, <50% present with classic signs and symptoms of pain in the right lower quadrant, mild fever and nausea. It is standard for patients who present with appendicitis to be managed operatively with a laparoscopic appendectomy within 24 hours, otherwise the risk of complications such as rupture, infection, and even death increases dramatically. CLINICAL FEATURES: This is a retrospective case report following a 27-year-old male with missed appendicitis, who presented to a chiropractor two-weeks after self-diagnosed food poisoning. On assessment, he was tender with resisted lumbar rotation. Psoas Sign, McBurney's Point, vascular exam, hip exam, were negative. A diagnosis of an abdominal strain was provided. Two weeks later, he returned to the chiropractor without an improvement of symptoms. INTERVENTION & OUTCOME: The patient was sent to the hospital, where he was provided a diagnosis of missed appendicitis. He required a hemicolonectomy due to the associated phlegmonous mass. SUMMARY: When a patient presents to a chiropractic clinic with symptoms of abdominal pain, having a comprehensive list of non-mechanical differential diagnoses as well as mechanical differentials is crucial. Appropriate assessment and management of abdominal cases decreases the risk to patients, as missed diagnoses often require more invasive interventions.

OBJECTIF: Cette étude de cas vise à souligner la nécessité d'une liste appropriée et détaillée de diagnostics différentiels lors de la gestion des patients, car il n'est pas suffisant de supposer que les cas sont d'ordre mécanique, jusqu'à la preuve du contraire. Il y a plus de 250 000 cas d'appendicite par an aux États-Unis. Parmi ces cas, < 50 % présentent des signes et des symptômes classiques de douleur dans le quadrant inférieur droit, de fièvre légère et de nausées. Il est normal qu'un patient qui se présente avec une appendicite soit géré par une intervention chirurgicale (appendicectomie par laparoscopie) dans les 24 heures, sinon le risque de complications, telles que rupture, infection et décès, augmente considérablement. CARACTÉRISTIQUES CLINIQUES: Ceci est une étude de cas rétrospective qui suit un homme de 27 ans dont le diagnostic d'appendicite a été manqué lorsqu'il s'est présenté à un chiropraticien deux semaines après un autodiagnostic d'intoxication alimentaire. Son examen avait révélé une sensibilité au toucher avec une résistance à la rotation lombaire. Le signe du psoas, le point de McBurney, l'examen vasculaire et l'examen de la hanche se sont révélés négatifs. Un diagnostic de claquage abdominal a été établi. Deux semaines plus tard, il est retourné au chiropraticien sans aucune amélioration des symptômes. INTERVENTION ET RÉSULTATS: Le patient a été envoyé à l'hôpital, où une appendicite manquée a été diagnostiquée. Il a fallu lui faire une hémicolectomie en raison de la masse phlegmoneuse associée. RÉSUMÉ: Quand un patient se présente à une clinique de chiropratique avec des symptômes de douleurs abdominales, il est crucial d'avoir une liste complète de diagnostics différentiels non-mécaniques ainsi que de différentiels mécaniques. L'évaluation et la gestion appropriées des douleurs abdominales diminuent le risque pour les patients, car les diagnostics manqués nécessitent souvent des interventions plus invasives.

Guyon Canal Syndrome: lack of management in a case of unresolved handlebar palsy.

OBJECTIVE: To present the clinical diagnostic features including management of Guyon canal syndrome in a case with unresolved sensory deficits in a young female cyclist. CLINICAL PRESENTATION: After 14 days of cycling across Canada, a 23-year old female experienced sensory loss, followed by atrophy and a "claw" hand appearance of her left hand. INTERVENTION AND OUTCOME: Treatment included cervical chiropractic manipulation, soft tissue therapy and the use of cycling gloves. Seven years after the initial injury a lack of sensation in the ulnar nerve distribution of her left hand has persisted. DISCUSSION: This case demonstrates that a lack of proper management can lead to permanent sensory loss and is worth highlighting. Various therapists evaluated the patient's symptoms and provided minimal care. No diagnosis was given, nor were appropriate measures taken for her to understand the risks of continuing to ride. SUMMARY: Although treatment for Guyon Canal Syndrome can be as easy as cessation from cycling until symptoms subside, other treatment options could be utilized to help manage ulnar nerve compression injuries in cyclists.

OBJECTIF: Présenter les caractéristiques de diagnostic clinique, dont le traitement du syndrome du canal de Guyon dans un cas de déficits sensoriels non traités chez une jeune cycliste. TABLEAU CLINIQUE: Après 14 jours de vélo à travers le Canada, une femme de 23 ans a souffert une perte sensorielle, suivie par une atrophie et l'aspect d'une « main en griffe ¼ à sa main gauche. INTERVENTION ET RÉSULTATS: Le traitement comportait la manipulation chiropratique cervicale, le traitement des tissus mous et le port de gants de vélo. Sept ans après la blessure initiale, un manque de sensation dans la distribution du nerf cubital de la main gauche a persisté. DISCUSSION: Ce cas démontre que le manque de traitement approprié peut entraîner la perte sensorielle permanente et mérite qu'on s'y attarde. Différents thérapeutes ont évalué les symptômes de la patiente et lui ont fourni des soins minimaux. Aucun diagnostic n'a été fait, ni des mesures appropriées n'étaient prises pour lui faire comprendre les risques de continuer à faire du vélo. RÉSUMÉ: Bien que le traitement du syndrome du canal de Guyon soit aussi simple que l'arrêt de la pratique du vélo jusqu'à la disparition des symptômes, d'autres options thérapeutiques pourraient être utilisées pour aider à traiter les lésions de compression du nerf cubital chez les cyclistes.

Consequences of the selective blockage of chaperone-mediated autophagy.

Chaperone-mediated autophagy (CMA) is a selective pathway for the degradation of cytosolic proteins in lysosomes. CMA declines with age because of a decrease in the levels of lysosome-associated membrane protein (LAMP) type 2A, a lysosomal receptor for this pathway. We have selectively blocked the expression of LAMP-2A in mouse fibroblasts in culture and analyzed the cellular consequences of reduced CMA activity. CMA-defective cells maintain normal rates of long-lived protein degradation by up-regulating macroautophagy, the major form of autophagy. Constitutive up-regulation of macroautophagy is unable, however, to compensate for all CMA functions. Thus, CMA-defective cells are more sensitive to stressors, suggesting that, although protein turnover is maintained, the selectivity of CMA is necessary as part of the cellular response to stress. Our results also denote the existence of cross-talk among different forms of autophagy.