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Nonlinear curve fitting is an important process in laboratory medicine, particularly with the increased use of highly sensitive antibody-based assays. Although the process is often automated in commercially available software, it is important that clinical scientists and physicians recognize the limitations of the various approaches used and are able to select the most appropriate model. This article summarizes the key nonlinear functions and demonstrates their application to common laboratory data. Following this, a basic overview of the statistical comparison of models is presented and then a discussion of important algorithms used in nonlinear curve fitting. An accompanying Microsoft Excel workbook is available that can be used to explore the content of this article.

Neurodegenerative diseases are amongst the most devastating of human disorders. New technologies have led to a rapid increase in the identification of disease-related genes with an enhanced appreciation of the key roles played by genetics in the etiology of these disorders. Importantly, pinpointing the normal function of disease gene proteins leads to new understanding of the cellular machineries and pathways that are altered in the disease process. One such emerging pathway is membrane trafficking in the endosomal system. This key cellular process controls the localization and levels of a myriad of proteins and is thus critical for normal cell function. In this review we will focus on three neurodegenerative diseases; Parkinson disease, amyotrophic lateral sclerosis, and hereditary spastic paraplegias, for which a large number of newly discovered disease genes encode proteins that function in endosomal membrane trafficking. We will describe how alterations in these proteins affect endosomal function and speculate on the contributions of these disruptions to disease pathophysiology.

Lysosomes help maintain cellular proteostasis, and defects in lysosomal positioning and function can cause disease, including neurodegenerative disorders. The spatiotemporal distribution of lysosomes is regulated by small GTPases including Rabs, which are activated by guanine nucleotide exchange factors (GEFs). DENN domain proteins are the largest family of Rab GEFs. Using a cell-based assay, we screened DENND6A, a member of the DENN domain protein family against all known Rabs and identified it as a potential GEF for 20 Rabs, including Rab34. Here, we demonstrate that DENND6A activates Rab34, which recruits a RILP/dynein complex to lysosomes, promoting lysosome retrograde transport. Further, we identify DENND6A as an effector of Arl8b, a major regulatory GTPase on lysosomes. We demonstrate that Arl8b recruits DENND6A to peripheral lysosomes to activate Rab34 and initiate retrograde transport, regulating nutrient-dependent lysosomal juxtanuclear repositioning. Loss of DENND6A impairs autophagic flux. Our findings support a model whereby Arl8b/DENND6A/Rab34-dependent lysosomal retrograde trafficking controls autophagy. Small GTPases such as Rabs control the positioning of lysosomes. Here, the authors unveil a molecular cascade orchestrated by Arl8/DENND6A/Rab34 that regulates lysosome location, impacting autophagy.

Background Post-deinstitutionalisation, mental health supported accommodation services have been implemented widely. The available research evidence is heterogeneous in nature and resistant to synthesis attempts, leaving researchers and policy makers with no clear summary what works and for whom. In this context, we undertook a comprehensive systematic review of quantitative studies in order to synthesise the current evidence on mental health and psychosocial outcomes for individuals residing in mental health supported accommodation services. Methods Using a combination of electronic database searches, hand searches, forward-backward snowballing and article recommendations from an expert panel, 115 papers were identified for review. Data extraction and quality assessments were conducted, and 33 articles were excluded due to low quality, leaving 82 papers in the final review. Variation in terminology and service characteristics made the comparison of service models unfeasible. As such, findings were presented according to the following sub-groups: ‘Homeless’, ‘Deinstitutionalisation’ and ‘General Severe Mental Illness (SMI)’. Results Results were mixed, reflecting the heterogeneity of the supported accommodation literature, in terms of research quality, experimental design, population, service types and outcomes assessed. There is some evidence that supported accommodation is effective across a range of psychosocial outcomes. The most robust evidence supports the effectiveness of the permanent supported accommodation model for homeless SMI in generating improvements in housing retention and stability, and appropriate use of clinical services over time, and for other forms of supported accommodation for deinstitutionalised populations in reducing hospitalisation rates and improving appropriate service use. The evidence base for general SMI populations is less developed, and requires further research. Conclusions A lack of high-quality experimental studies, definitional inconsistency and poor reporting continue to stymie our ability to identify effective supported accommodation models and practices. The authors recommend improved reporting standards and the prioritisation of experimental studies that compare outcomes across different service models.

Understanding Alzheimer's disease (AD) etiology requires a comprehensive molecular and biological investigation of the estimated 184 genes/proteins linked to AD risk. However, most AD research focuses on 3 genes, APP, APOE, and TAU, which receive 49% of all AD-related PubMed hits. This highlights the need for a broader exploration of AD risk factors. A major challenge in this regard is the limited availability of selective and renewable antibodies for studying all AD-associated proteins. The Target Enablement to Accelerate Therapy Development for Alzheimer's Disease (TREAT-AD) program focuses on developing or identifying high-quality research tools for less-studied AD-associated proteins, enabling hypothesis testing (Axtman et al., Alzheimers Dement, 2023). Antibody Characterization through Open Science (YCharOS) is an international, public-good initiative that unites academic researchers, funding agencies, leading antibody manufacturers and knockout cell line providers to evaluate antibody performance (Ayoubi et al., Nature Protocols, 2024). Using knockout cell lines as isogenic controls, antibodies from multiple manufacturers are tested side-by-side in western blot, immunoprecipitation and immunofluorescence. Results are disseminated openly and rapidly via the AD Knowledge Portal (https://adknowledgeportal.synapse.org/). To date, 441 antibodies targeting 41 community-prioritized AD targets have been characterized. Most cited antibodies for these proteins are polyclonal and often lack specificity, particularly in immunofluorescence experiments. However, our findings reveal that selective and renewable antibodies, many of which are newly generated and rarely used in published studies, were available for 90% of TREAT-AD targets. When all existing antibodies for a protein underperform, TREAT-AD generates recombinant proteins as antigens for antibody development, collaborating with YCharOS partners for antibody production. This approach has been applied to AD-related proteins, including SMOC1, yielding recombinant antibodies with superior selectivity over existing ones. Using knockout-validated antibodies, we demonstrated that the proteins CD44, Moesin, sFRP-1 and Midkine, identified through systems biology by the TREAT-AD initiative as having altered gene expression in AD, are upregulated under disease conditions in AD mouse models (Doolen et al. F1000Research, 2024). YCharOS has established characterization standards that align with funding and journal requirements, and is committed to identify reliable and accessible tools for AD research.

Letter to the Editor

AP-2 is the core-organizing element in clathrin-mediated endocytosis. During the formation of clathrin-coated vesicles, clathrin and endocytic accessory proteins interact with AP-2 in a temporally and spatially controlled manner, yet it remains elusive as to how these interactions are regulated. Here, we demonstrate that the endocytic protein NECAP 1, which binds to the α-ear of AP-2 through a C-terminal WxxF motif, uses an N-terminal PH-like domain to compete with clathrin for access to the AP-2 β2-linker, revealing a means to allow AP-2-mediated coordination of accessory protein recruitment and clathrin polymerization at sites of vesicle formation. Knockdown and functional rescue studies demonstrate that through these interactions, NECAP 1 and AP-2 cooperate to increase the probability of clathrin-coated vesicle formation and to control the number, size, and cargo content of the vesicles. Together, our data demonstrate that NECAP 1 modulates the AP-2 interactome and reveal a new layer of organizational control within the endocytic machinery.

Virtually all prospective medical students are required to take courses in general and organic chemistry, but are they really necessary? We discuss the relevance of chemistry to modern medicine and the arguments for and against its use as an essential prerequisite.

Abstract Objective Assays based on redox reactions that involve proton transfer are vulnerable to artifactual findings in metabolic acidosis/alkalosis. We evaluated the impact of pH on the measurement of blood glucose by the glucose dehydrogenase/pyrroloquinoline quinone system used in point-of-care-testing. Methods We applied a series of thermodynamic equations to adjust the Gibbs energy for the pyrroloquinoline quinone couple. This adjusts values taken under standard conditions to those more closely resembling the physiological state. Results Under standard conditions, the pyrroloquinoline quinone couple has Eo = −0.125 V whereas adjustment to the physiological state (pH 7.40, ionic strength 0.15 mol/L, and temperature 310.15°K) yields Eo′ = −0.166 V. This corresponds to an uncertainty in blood glucose determination of approximately 0.13 mmol/L. Conclusion We have demonstrated that the impact of pH on blood glucose determination by the glucose dehydrogenase/pyrroloquinoline quinone system (under physiologically relevant conditions of ionic strength and temperature) is not clinically significant.

Lewy bodies (LBs), α-synuclein-enriched intracellular inclusions, are a hallmark of Parkinson’s disease (PD) pathology, yet a cellular model for LB formation remains elusive. Recent evidence indicates that immune dysfunction may contribute to the development of PD. In this study, we found that induced pluripotent stem cell (iPSC)-derived human dopaminergic (DA) neurons form LB-like inclusions after treatment with α-synuclein preformed fibrils (PFFs) but only when coupled to a model of immune challenge (interferon-γ or interleukin-1β treatment) or when co-cultured with activated microglia-like cells. Exposure to interferon-γ impairs lysosome function in DA neurons, contributing to LB formation. The knockdown of LAMP2 or the knockout of GBA in conjunction with PFF administration is sufficient for inclusion formation. Finally, we observed that the LB-like inclusions in iPSC-derived DA neurons are membrane bound, suggesting that they are not limited to the cytoplasmic compartment but may be formed due to dysfunctions in autophagy. Together, these data indicate that immune-triggered lysosomal dysfunction may contribute to the development of PD pathology. Bayati et al. discovered that sequential treatment of iPSC-derived dopaminergic neurons with α-synuclein fibrils and proinflammatory cytokines leads to the formation of Lewy body–like inclusions, through the downregulation of lysosomal proteins.