An organelle-specific protein landscape identifies novel diseases and molecular mechanisms

Cellular organelles provide opportunities to relate biological mechanisms to disease. Here we use affinity proteomics, genetics and cell biology to interrogate cilia: poorly understood organelles, where defects cause genetic diseases. Two hundred and seventeen tagged human ciliary proteins create a final landscape of 1,319 proteins, 4,905 interactions and 52 complexes. Reverse tagging, repetition of purifications and statistical analyses, produce a high-resolution network that reveals organelle-specific interactions and complexes not apparent in larger studies, and links vesicle transport, the cytoskeleton, signalling and ubiquitination to ciliary signalling and proteostasis. We observe sub-complexes in exocyst and intraflagellar transport complexes, which we validate biochemically, and by probing structurally predicted, disruptive, genetic variants from ciliary disease patients. The landscape suggests other genetic diseases could be ciliary including 3M syndrome. We show that 3M genes are involved in ciliogenesis, and that patient fibroblasts lack cilia. Overall, this organelle-specific targeting strategy shows considerable promise for Systems Medicine

Nationwide Association of Surgical Performance of Minimally Invasive Esophagectomy With Patient Outcomes

IMPORTANCE: Suboptimal surgical performance is hypothesized to be associated with less favorable patient outcomes in minimally invasive esophagectomy (MIE). Establishing this association may lead to programs that promote better surgical performance of MIE and improve patient outcomes.OBJECTIVE: To investigate associations between surgical performance and postoperative outcomes after MIE.DESIGN, SETTING, AND PARTICIPANTS: In this nationwide cohort study of 15 Dutch hospitals that perform more than 20 MIEs per year, 7 masked expert MIE surgeons assessed surgical performance using videos and a previously developed and validated competency assessment tool (CAT). Each hospital submitted 2 representative videos of MIEs performed between November 4, 2021, and September 13, 2022. Patients registered in the Dutch Upper Gastrointestinal Cancer Audit between January 1, 2020, and December 31, 2021, were included to examine patient outcomes.EXPOSURE: Hospitals were divided into quartiles based on their MIE-CAT performance score. Outcomes were compared between highest (top 25%) and lowest (bottom 25%) performing quartiles. Transthoracic MIE with gastric tube reconstruction.MAIN OUTCOME AND MEASURE: The primary outcome was severe postoperative complications (Clavien-Dindo ≥3) within 30 days after surgery. Multilevel logistic regression, with clustering of patients within hospitals, was used to analyze associations between performance and outcomes.RESULTS:In total, 30 videos and 970 patients (mean [SD] age, 66.6 [9.1] years; 719 men [74.1%]) were included. The mean (SD) MIE-CAT score was 113.6 (5.5) in the highest performance quartile vs 94.1 (5.9) in the lowest. Severe postoperative complications occurred in 18.7% (41 of 219) of patients in the highest performance quartile vs 39.2% (40 of 102) in the lowest (risk ratio [RR], 0.50; 95% CI, 0.24-0.99). The highest vs the lowest performance quartile showed lower rates of conversions (1.8% vs 8.9%; RR, 0.21; 95% CI, 0.21-0.21), intraoperative complications (2.7% vs 7.8%; RR, 0.21; 95% CI, 0.04-0.94), and overall postoperative complications (46.1% vs 65.7%; RR, 0.54; 95% CI, 0.24-0.96). The R0 resection rate (96.8% vs 94.2%; RR, 1.03; 95% CI, 0.97-1.05) and lymph node yield (mean [SD], 38.9 [14.7] vs 26.2 [9.0]; RR, 3.20; 95% CI, 0.27-3.21) increased with oncologic-specific performance (eg, hiatus dissection, lymph node dissection). In addition, a high anastomotic phase score was associated with a lower anastomotic leakage rate (4.6% vs 17.7%; RR, 0.14; 95% CI, 0.06-0.31).CONCLUSIONS AND RELEVANCE: These findings suggest that better surgical performance is associated with fewer perioperative complications for patients with esophageal cancer on a national level. If surgical performance of MIE can be improved with MIE-CAT implementation, substantially better patient outcomes may be achievable.</p

Developments in cell biology for quantitative immunoelectron microscopy based on thin sections: a review

Quantitative immunoelectron microscopy uses ultrathin sections and gold particle labelling to determine distributions of molecules across cell compartments. Here, we review a portfolio of new methods for comparing labelling distributions between different compartments in one study group (method 1) and between the same compartments in two or more groups (method 2). Specimen samples are selected unbiasedly and then observed and expected distributions of gold particles are estimated and compared by appropriate statistical procedures. The methods can be used to analyse gold label distributed between volume-occupying (organelle) and surface-occupying (membrane) compartments, but in method 1, membranes must be treated as organelles. With method 1, gold counts are combined with stereological estimators of compartment size to determine labelling density (LD). For volume-occupiers, LD can be expressed simply as golds per test point and, for surface-occupiers, as golds per test line intersection. Expected distributions are generated by randomly assigning gold particles to compartments and expressing observed/expected counts as a relative labelling index (RLI). Preferentially-labelled compartments are identified from their RLI values and by Chi-squared analysis of observed and expected distributions. For method 2, the raw gold particle counts distributed between compartments are simply compared across groups by contingency table and Chi-squared analysis. This identifies the main compartments responsible for the differences between group distributions. Finally, we discuss labelling efficiency (the number of gold particles per target molecule) and describe how it can be estimated for volume- or surface-occupiers by combining stereological data with biochemical determinations

Plasmodium falciparum: Differential Selection of Drug Resistance Alleles in Contiguous Urban and Peri-Urban Areas of Brazzaville, Republic of Congo

The African continent is currently experiencing rapid population growth, with rising urbanization increasing the percentage of the population living in large towns and cities. We studied the impact of the degree of urbanization on the population genetics of Plasmodium falciparum in urban and peri-urban areas in and around the city of Brazzaville, Republic of Congo. This field setting, which incorporates local health centers situated in areas of varying urbanization, is of interest as it allows the characterization of malaria parasites from areas where the human, parasite, and mosquito populations are shared, but where differences in the degree of urbanization (leading to dramatic differences in transmission intensity) cause the pattern of malaria transmission to differ greatly. We have investigated how these differences in transmission intensity affect parasite genetic diversity, including the amount of genetic polymorphism in each area, the degree of linkage disequilibrium within the populations, and the prevalence and frequency of drug resistance markers. To determine parasite population structure, heterozygosity and linkage disequilibrium, we typed eight microsatellite markers and performed haplotype analysis of the msp1 gene by PCR. Mutations known to be associated with resistance to the antimalarial drugs chloroquine and pyrimethamine were determined by sequencing the relevant portions of the crt and dhfr genes, respectively. We found that parasite genetic diversity was comparable between the two sites, with high levels of polymorphism being maintained in both areas despite dramatic differences in transmission intensity. Crucially, we found that the frequencies of genetic markers of drug resistance against pyrimethamine and chloroquine differed significantly between the sites, indicative of differing selection pressures in the two areas

The design of an optimal Bonus-Malus System based on the Sichel distribution

This chapter presents the design of an optimal Bonus-Malus System (BMS) using the Sichel distribution to model the claim frequency distribution. This system is proposed as an alternative to the optimal BMS obtained by the traditional Negative Binomial model [19]. The Sichel distribution has a thicker tail than the Negative Binomial distribution and it is considered as a plausible model for highly dispersed count data. We also consider the optimal BMS provided by the Poisson-Inverse Gaussian distribution (PIG), which is a special case of the Sichel distribution. Furthermore, we develop a generalised BMS that takes into account both the a priori and a posteriori characteristics of each policyholder. For this purpose we consider the generalised additive models for location, scale and shape (GAMLSS) in order to use all available information in the estimation of the claim frequency distribution. Within the framework of the GAMLSS we propose the Sichel GAMLSS for assessing claim frequency as an alternative to the Negative Binomial Type I (NBI) regression model used by Dionne and Vanasse [9, 10]. We also consider the NBI and PIG GAMLSS for assessing claim frequency