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Autophagy, a self-defense mechanism, has been found to be associated with drug resistance in hepatocellular carcinoma (HCC). Our study was designed to investigate the role and related mechanisms of autophagy in matrine-induced apoptosis in hepatoma cells of HepG2 and Bel7402. Cell apoptosis was detected by flow cytometry analysis (Annexin V–FITC/PI double-staining assay), the activity and activating cleavages of caspase-3, -8, and -9. MTT assay and colony forming assay were used to assess the effect of matrine on growth and proliferation of HCC cells. Autophagic flux in HCC cells was analyzed using the expression of LC3BI/II and p62/SQSTM1, GFP-LC3 transfection, and transmission electron microscopy. Moreover, regarding to the associated mechanisms, the effects of matrine on the phosphoinositide 3-kinase/AKT/mTOR pathway and beclin-1 were studied. Our results showed that: (1) both autophagy and apoptosis could be induced by treatment with matrine; (2) using the autophagic inhibitor chloroquine and beclin-1 small-interfering RNA, cell apoptosis induced by matrine could be enhanced in a caspase-dependent manner; and (3) autophagy was induced via inhibition of PI3K/AKT/mTOR pathway and up-regulation of beclin-1. In conclusion, inhibition of autophagy could enhance matrine-induced apoptosis in human hepatoma cells.

ObjectiveTo evaluate whether the clinical benefit and relapse rates in anti-muscle-specific kinase (MuSK) myasthenia gravis (MG) differ depending on the protocol of rituximab followed. MethodsThis retrospective multicentre study in patients with MuSK MG compared three rituximab protocols in terms of clinical status, relapse, changes in treatment, and adverse side effects. The primary effectiveness endpoint was clinical relapse requiring a further infusion of rituximab. Survival curves were estimated using Kaplan-Meier methods and survival analyses were undertaken using Cox proportional-hazards models. ResultsTwenty-five patients were included: 11 treated with protocol 4 + 2 (375 mg/m(2)/4 weeks, then monthly for 2 months), five treated with protocol 1 + 1 (two 1 g doses 2 weeks apart), and nine treated with protocol 4 (375 mg/m(2)/4 weeks). Mean follow-up was 5.0 years (SD 3.3). Relapse occurred in 18.2%, 80%, and 33.3%, and mean time to relapse was 3.5 (SD 1.5), 1.1 (SD 0.4), and 2.5 (SD 1.4) years, respectively. Based on Kaplan-Meier estimates, patients treated with protocol 4 + 2 had fewer and later relapses than patients treated with the other two protocols (log-rank test P = 0.0001). Patients treated with protocol 1 + 1 had a higher risk of relapse than patients treated with protocol 4 + 2 (HR 112.8, 95% CI, 5.7-2250.4, P = 0.002). Patients treated with protocol 4 showed a trend to a higher risk of relapse than those treated with protocol 4 + 2 (HR 9.2, 95% CI 0.9-91.8, P = 0.059). InterpretationThis study provides class IV evidence that the 4 + 2 rituximab protocol has a lower clinical relapse rate and produces a more durable response than the 1 + 1 and 4 protocols in patients with MuSK MG.

AbstractThere is now widespread acceptance of the universal coverage approach, presented in the 2010 World Health Report. There are more and more voices for the benefit of creating a single national risk pool. Now, a body of literature is emerging on institutional design and organizational practice for universal coverage, related to management of the three health-financing functions: collection, pooling and purchasing. While all countries can move towards universal coverage, lower-income countries face particular challenges, including scarce resources and limited capacity. Recently, the Lao PDR has been preparing options for moving to a single national health insurance scheme. The aim is to combine four different social health protection schemes into a national health insurance authority (NHIA) with a single national fund- and risk-pool. This paper investigates the main institutional and organizational challenges related to the creation of the NHIA. The paper uses a qualitative approach, drawing on the World Health Organization's institutional and Organizational Assessment for Improving and Strengthening health financing (OASIS) conceptual framework for data analysis. Data were collected from a review of key health financing policy documents and from 17 semi-structured key informant interviews. Policy makers and advisors are confronting issues related to institutional arrangements, funding sources for the authority and government support for subsidies to the demand-side health financing schemes. Compulsory membership is proposed, but the means for covering the informal sector have not been resolved. While unification of existing schemes may be the basis for creating a single risk pool, challenges related to administrative capacity and cross-subsidies remain. The example of Lao PDR illustrates the need to include consideration of national context, the sequencing of reforms and the time-scale appropriate for achieving universal coverage.

Cryo-electron microscopy (cryo-EM) is a powerful technique for determining the structure of proteins and other macromolecular complexes at near-atomic resolution. In single particle cryo-EM, the central problem is to reconstruct the three-dimensional structure of a macromolecule from $10^{4-7}$ noisy and randomly oriented two-dimensional projections. However, the imaged protein complexes may exhibit structural variability, which complicates reconstruction and is typically addressed using discrete clustering approaches that fail to capture the full range of protein dynamics. Here, we introduce a novel method for cryo-EM reconstruction that extends naturally to modeling continuous generative factors of structural heterogeneity. This method encodes structures in Fourier space using coordinate-based deep neural networks, and trains these networks from unlabeled 2D cryo-EM images by combining exact inference over image orientation with variational inference for structural heterogeneity. We demonstrate that the proposed method, termed cryoDRGN, can perform ab initio reconstruction of 3D protein complexes from simulated and real 2D cryo-EM image data. To our knowledge, cryoDRGN is the first neural network-based approach for cryo-EM reconstruction and the first end-to-end method for directly reconstructing continuous ensembles of protein structures from cryo-EM images.

AbstractOnly a few studies have assessed the long-term duration of the humoral immune response against severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2).In this nationwide longitudinal study from the Faroe Islands with close to full participation of all individuals on the Islands with PCR confirmed COVID-19 during the two waves of infections in the spring and autumn 2020 (n=172 & n=233), samples were drawn at three longitudinal time points (3, 7 and 12 months and 1, 3 and 7 months after disease onset, respectively).Serum was analyzed with a direct quantitative IgG antibody binding ELISA to detect anti–SARS-CoV-2 spike RBD antibodies and a commercially available qualitative sandwich RBD ELISA kit measuring total antibody binding.The seropositive rate in the convalescent individuals was above 95 % at all sampling time points for both assays. There was an overall decline in IgG titers over time in both waves (p < 0.001). Pairwise comparison showed that IgG declined significantly from the first sample until approximately 7 months in both waves (p < 0.001). After that, the antibody level still declined significantly (p < 0.001), but decelerated with an altered slope remaining fairly stable from 7 months to 12 months after infection. Interestingly, the IgG titers followed a U-shaped curve with higher antibody levels among the oldest (67+) and the youngest (0– 17) age groups compared to intermediate groups (p < 0.001).Our results indicate that COVID-19 convalescent individuals are likely to be protected from reinfection up to 12 months after symptom onset and maybe even longer. We believe our results can add to the understanding of natural immunity and the expected durability of SARS-CoV-2 vaccine immune responses.

One of the main hypotheses in fine-grained complexity is that All-Pairs Shortest Paths (APSP) for $n$-node graphs requires $n^{3-o(1)}$ time. Another famous hypothesis is that the $3$SUM problem for $n$ integers requires $n^{2-o(1)}$ time. Although there are no direct reductions between $3$SUM and APSP, it is known that they are related: there is a problem, $(\min,+)$-convolution that reduces in a fine-grained way to both, and a problem Exact Triangle that both fine-grained reduce to. In this paper we find more relationships between these two problems and other basic problems. P\u{a}tra\c{s}cu had shown that under the $3$SUM hypothesis the All-Edges Sparse Triangle problem in $m$-edge graphs requires $m^{4/3-o(1)}$ time. The latter problem asks to determine for every edge $e$, whether $e$ is in a triangle. It is equivalent to the problem of listing $m$ triangles in an $m$-edge graph where $m=\tilde{O}(n^{1.5})$, and can be solved in $O(m^{1.41})$ time [Alon et al.'97] with the current matrix multiplication bounds, and in $\tilde{O}(m^{4/3})$ time if $\omega=2$. We show that one can reduce Exact Triangle to All-Edges Sparse Triangle, showing that All-Edges Sparse Triangle (and hence Triangle Listing) requires $m^{4/3-o(1)}$ time also assuming the APSP hypothesis. This allows us to provide APSP-hardness for many dynamic problems that were previously known to be hard under the $3$SUM hypothesis. We also consider the previously studied All-Edges Monochromatic Triangle problem. Via work of [Lincoln et al.'20], our result on All-Edges Sparse Triangle implies that if the All-Edges Monochromatic Triangle problem has an $O(n^{2.5-\epsilon})$ time algorithm for $\epsilon>0$, then both the APSP and $3$SUM hypotheses are false. We also connect the problem to other ``intermediate'' problems, whose runtimes are between $O(n^\omega)$ and $O(n^3)$, such as the Max-Min product problem. Comment: To appear in FOCS'20

Encouraged by the success of deep neural networks on a variety of visual tasks, much theoretical and experimental work has been aimed at understanding and interpreting how vision networks operate. Meanwhile, deep neural networks have also achieved impressive performance in audio processing applications, both as sub-components of larger systems and as complete end-to-end systems by themselves. Despite their empirical successes, comparatively little is understood about how these audio models accomplish these tasks. In this work, we employ a recently developed statistical mechanical theory that connects geometric properties of network representations and the separability of classes to probe how information is untangled within neural networks trained to recognize speech. We observe that speaker-specific nuisance variations are discarded by the network's hierarchy, whereas task-relevant properties such as words and phonemes are untangled in later layers. Higher level concepts such as parts-of-speech and context dependence also emerge in the later layers of the network. Finally, we find that the deep representations carry out significant temporal untangling by efficiently extracting task-relevant features at each time step of the computation. Taken together, these findings shed light on how deep auditory models process time dependent input signals to achieve invariant speech recognition, and show how different concepts emerge through the layers of the network. Comment: Advances in Neural Information Processing Systems. 2019

ABSTRACT In Lao People’s Democratic Republic (Lao PDR), reports on disease frequency are very limited. This study aimed to report frequencies of the main cause of admission among inpatients of a tertiary general hospital (Mittaphab Hospital) in Vientiane. Subjects were inpatients who were admitted from January 3 to February 2 in 2017. The dataset were made as a pilot run to establish hospital statistics. The data on sex, age, address (province), dates of admission and discharge, and main diagnosis were collected from paper-based medical charts. International Classification of Diseases 10 was applied for classifying the main diagnosis. During the 1-month period, 1,201 inpatients (637 males and 564 females) were admitted, including 171 (14.2%) aged <20 years and 254 (21.1%) aged ≥60 years. About 20% patients were from outside of Vientiane. Among them, 67.5% (62.5% in males and 73.8% in females) were admitted within 7 days. The main causes with more than 10% in males were injury and poisoning S00-T98 (49.8%), while those in females were injury and poisoning S00-T98 (25.2%), pregnancy and childbirth O00-O99 (19.0%), and diseases of genitourinary system N00-N99 (13.7%). Injury and poisoning S00-T98 among inpatients aged <20 years was 81.8% in males and 59.0% in females. Among those aged 20–59 years, it was 49.9% and 22.4%, and among those aged ≥60 years it was 22.3% and 16.9%, respectively. This is the first report on the frequencies of main diseases among inpatients in Lao PDR. Injury was the first main cause of admission at the tertiary hospital.

Abstract Background Testing for HIV at birth has the potential to identify infants infected in utero, and allows for the possibility of beginning treatment immediately after birth; point of care (POC) testing allows rapid return of results and faster initiation on treatment for positive infants. Eswatini piloted birth testing in three public maternities for over 2 years. Methods In order to assess the acceptability of POC birth testing in the pilot sites in Eswatini, interviews were held with caregivers of HIV-exposed infants who were offered birth testing (N = 28), health care workers (N = 14), and policymakers (N = 10). Participants were purposively sampled. Interviews were held in English or SiSwati, and transcribed in English. Transcripts were coded by line, and content analysis and constant comparison were used to identify key themes for each respondent type. Results Responses were categorized into: knowledge, experience, opinions, barriers and challenges, facilitators, and suggestions to improve POC birth testing. Preliminary findings reveal that point of care birth testing has been very well received but challenges were raised. Most caregivers appreciated testing the newborns at birth and getting results quickly, since it reduced anxiety of waiting for several weeks. However, having a favorable experience with testing was linked to having supportive and informed family members and receiving a negative result. Caregivers did not fully understand the need for blood draws as opposed to tests with saliva, and expressed the fears of seeing their newborns in pain. They were specifically grateful for supportive nursing staff who respected their confidentiality. Health care workers expressed strong support for the program but commented on the high demand for testing, increased workload, difficulty with errors in the testing machine itself, and struggles to implement the program without sufficient staffing, especially on evenings and weekends when phlebotomists were not available. Policymakers noted that there have been challenges within the program of losing mothers to follow up after they leave hospital, and recommended stronger linkages to community groups. Conclusions There is strong support for scale-up of POC birth testing, but countries should consider ways to optimize staffing and manage demand.