Single-cell transcriptomes of the regenerating intestine reveal a revival stem cell.

The turnover of the intestinal epithelium is driven by multipotent LGR5+ crypt-base columnar cells (CBCs) located at the bottom of crypt zones1. However, CBCs are lost following injury, such as irradiation2, but the intestinal epithelium is nevertheless able to recover3. Thus, a second population of quiescent '+4' cells, or reserve stem cells (RSCs), has previously been proposed to regenerate the damaged intestine4-7. Although CBCs and RSCs were thought to be mutually exclusive4,8, subsequent studies have found that LGR5+ CBCs express RSC markers9 and that RSCs were dispensable-whereas LGR5+ cells were essential-for repair of the damaged intestine3. In addition, progenitors of absorptive enterocytes10, secretory cells11-15 and slow cycling LGR5+ cells16 have been shown to contribute to regeneration whereas the transcriptional regulator YAP1, which is important for intestinal regeneration, was suggested to induce a pro-survival phenotype in LGR5+ cells17. Thus, whether cellular plasticity or distinct cell populations are critical for intestinal regeneration remains unknown. Here we applied single-cell RNA sequencing to profile the regenerating mouse intestine and identified a distinct, damage-induced quiescent cell type that we term the revival stem cell (revSC). revSCs are marked by high clusterin expression and are extremely rare under homoeostatic conditions, yet give rise-in a temporal hierarchy-to all the major cell types of the intestine, including LGR5+ CBCs. After intestinal damage by irradiation, targeted ablation of LGR5+ CBCs, or treatment with dextran sodium sulfate, revSCs undergo a YAP1-dependent transient expansion, reconstitute the LGR5+ CBC compartment and are required to regenerate a functional intestine. These studies thus define a unique stem cell that is mobilized by damage to revive the homoeostatic stem cell compartment and regenerate the intestinal epithelium.

The Artefactual Branch Effect and Phylogenetic Conflict: Species Delimitation with Gene Flow in Mangrove Pit Vipers (Trimeresurus purpureomaculatus-erythrurus Complex).

Mangrove pit vipers of the Trimeresurus purpureomaculatus-erythrurus complex are the only species of viper known to naturally inhabit mangroves. Despite serving integral ecological functions in mangrove ecosystems, the evolutionary history, distribution, and species boundaries of mangrove pit vipers remain poorly understood, partly due to overlapping distributions, confusing phenotypic variations, and the lack of focused studies. Here, we present the first genomic study on mangrove pit vipers and introduce a robust hypothesis-driven species delimitation framework that considers gene flow and phylogenetic uncertainty in conjunction with a novel application of a new class of speciation-based delimitation model implemented through the program Delineate. Our results showed that gene flow produced phylogenetic conflict in our focal species and substantiates the artefactual branch effect where highly admixed populations appear as divergent nonmonophyletic lineages arranged in a stepwise manner at the basal position of clades. Despite the confounding effects of gene flow, we were able to obtain unequivocal support for the recognition of a new species based on the intersection and congruence of multiple lines of evidence. This study demonstrates that an integrative hypothesis-driven approach predicated on the consideration of multiple plausible evolutionary histories, population structure/differentiation, gene flow, and the implementation of a speciation-based delimitation model can effectively delimit species in the presence of gene flow and phylogenetic conflict.

The effectiveness of physiotherapy interventions on fecal incontinence and quality of life following colorectal surgery: a systematic review and meta-analysis of randomized controlled trials.

PURPOSE: To investigate the effectiveness of physiotherapy interventions compared to control conditions on fecal incontinence (FI) and quality of life (QoL) following colorectal surgery. METHODS: Electronic searches in English-language (Scopus, Web of Science, Embase, AMED, CENTRAL, CINAHL, MEDLINE, Ovid, and PEDro) and Chinese-language (CNKI, Wanfang Data) databases were conducted. Trials comparing physiotherapy interventions against control conditions and assessing FI and QoL outcomes were included in the review. RESULTS: Ten trials were included. Meta-analysis revealed statistically significant improvements in lifestyle (0.54; 95% CI 0.03, 1.05; p = 0.04), coping behavior (MD 1.136; 95% CI 0.24, 2.04; p = 0.01), and embarrassment (0.417; 95% CI 0.14, 0.70; p = 0.00) components of QoL among individuals receiving pelvic floor muscle training (PFMT) compared with those receiving usual care (UC). Meta-analysis showed biofeedback to be significantly more effective than UC in enhancing anal resting pressure (ARP; 9.551; 95% CI 2.60, 16.51; p = 0.007), maximum squeeze pressure (MSP; 25.29; 95% CI 4.08, 48.50; p = 0.02), and rectal resting pressure (RRP; 0.51; 95% CI 0.10, 0.9; p = 0.02). Meta-analysis also found PFMT combined with biofeedback to be significantly more effective than PFMT alone for ARP (3.00; 95% CI 0.40, 5.60; p = 0.02), MSP (9.35, 95% CI 0.17, 18.53; p = 0.05), and RRP (1.54; 95% CI 0.60, 2.47; p = 0.00). CONCLUSIONS: PFMT combined with biofeedback was more effective than PFMT alone, but both interventions delivered alone were superior to UC. Future studies remain necessary to optimize and standardize the PFMT parameters for improving QoL among individuals who experience FI following CRC surgery. REVIEW REGISTRATION: This systematic review is registered in the PROSPERO registry (Ref: CRD42022337084).

Parastomal hernia repairs: A nationwide cohort study in the Republic of Ireland.

BACKGROUND: In the context of improving colorectal cancer outcomes, post-survivorship quality of life has become an important outcome measure. Parastomal hernias and their associated morbidity remain largely under-reported and under-appreciated. Despite their burden, conservative management is common. This study aims to provide a national overview on the current trends in parastomal hernia repairs (PHRs). METHODS: All PHRs performed in public hospitals across the country between 1/2017 to 7/2022 were identified retrospectively from the National Quality Assurance and Improvement System (NQAIS) database. Anonymised patient characteristics and quality indices were extracted for statistical analysis. RESULTS: A total of 565 PHRs, 64.1 % elective and the remainder emergent, were identified across 27 hospitals. The 8 national colorectal units performed 67.3 % of all repairs. While 42.3 % of PHRs were standalone procedures, reversal of Hartmann's procedure was the commonest simultaneous procedure in the remainder. The median age, ASA and Charlson Co-Morbidity Index were 64 years (19), 3(1) and 3(10) respectively. Mean length of stay (LOS) was 16.25 days (SD = 29.84). Linear regression analysis associated ASA (95 % CI 0.58-16.08, p < 0.035) and emergency admissions (95 % CI 5.86-25.55, P < 0.002) with a significantly longer LOS, with the latter also associated with more frequent emergency re-admissions (95 % CI 0.18-0.82, p < 0.002). CONCLUSION: Patients undergoing emergency PHR were older and significantly more comorbid. Consequently, these patients were subjected to longer hospital stays, more frequent readmissions and overall higher hospital costs. Multidisciplinary perioperative optimisation and standardised referral pathways should underpin the shift towards elective PHRs.

Clusters of Multiple Mutations: Incidence and Molecular Mechanisms.

It has been long understood that mutation distribution is not completely random across genomic space and in time. Indeed, recent surprising discoveries identified multiple simultaneous mutations occurring in tiny regions within chromosomes while the rest of the genome remains relatively mutation-free. Mechanistic elucidation of these phenomena, called mutation showers, mutation clusters, or kataegis, in parallel with findings of abundant clustered mutagenesis in cancer genomes, is ongoing. So far, the combination of factors most important for clustered mutagenesis is the induction of DNA lesions within unusually long and persistent single-strand DNA intermediates. In addition to being a fascinating phenomenon, clustered mutagenesis also became an indispensable tool for identifying a previously unrecognized major source of mutation in cancer, APOBEC cytidine deaminases. Future research on clustered mutagenesis may shed light onto important mechanistic details of genome maintenance, with potentially profound implications for human health.

Correction: Repair of multiple simultaneous double-strand breaks causes bursts of genome-wide clustered hypermutation.

[This corrects the article DOI: 10.1371/journal.pbio.3000464.].

C-terminal modification of a de novo designed antimicrobial peptide via capping of macrolactam rings.

In this work, by capping a macrolactam ring at the C-terminus of a de novo-designed peptide, namely zp80, we have constructed a small peptide library via the solid phase peptide synthesis for screening. Eight peptides bearing different aspartic acid-rich macrolactam rings but the same linear (IIRR)4 unit exhibited improved antibacterial activities, hemolytic activity, and selectivity index. Mechanistic studies revealed that they could destroy the integrity of bacterial envelope, leading to cytoplasm leakage and rapid dissipation of membrane potential. One of these peptides, zp90 with a macrolactam ring of (KaDGD), demonstrated preferential interaction with calcium ions at a stoichiometric ratio of 1:1, promoting the affinity of designed peptides to bacterial membrane. Overall, this work provides a feasible strategy for medicinal chemists to further develop potent, selective, and multifunctional de novo-designed antimicrobial peptides.

A novel combination treatment of antiADAM17 antibody and erlotinib to overcome acquired drug resistance in non-small cell lung cancer through the FOXO3a/FOXM1 axis.

After the identification of specific epidermal growth factor receptor (EGFR)-activating mutations as one of the most common oncogenic driver mutations in non-small cell lung cancer (NSCLC), several EGFR-tyrosine kinase inhibitors (EGFR-TKIs) with different clinical efficacies have been approved by various health authorities in the last two decades in targeting NSCLC harboring specific EGFR-activating mutations. However, most patients whose tumor initially responded to the first-generation EGFR-TKI developed acquired resistance. In this study, we developed a novel combination strategy, "antiADAM17 antibody A9(B8) + EGFR-TKIs", to enhance the efficacy of EGFR-TKIs. The addition of A9(B8) was shown to restore the effectiveness of erlotinib and overcome acquired resistance. We found that when A9(B8) antibody was treated with erlotinib or gefitinib, the combination treatment synergistically increased apoptosis in an NSCLC cell line and inhibited tumor growth in vivo. Interestingly, the addition of A9(B8) could only reduce the survival of the erlotinib-resistant NSCLC cell line and inhibit the growth of erlotinib-resistant tumors in vivo but not gefitinib-resistant cells. Furthermore, we revealed that A9(B8) overcame erlotinib resistance through the FOXO3a/FOXM1 axis and arrested the cell cycle at the G1/S phase, resulting in the apoptosis of cancer cells. Hence, this study establishes a novel, promising strategy for overcoming acquired resistance to erlotinib through the FOXO3a/FOXM1 axis by arresting the cell cycle at the G1/S phase.

Opioid prescribing after breast surgery: A systematic review of guidelines.

BACKGROUND: Despite advances in opioid-sparing analgesia, opioid prescribing in breast surgery remains suboptimal. Besides delayed rehabilitation, excess post-operative opioids may contribute significantly to opioid dependence. This systematic review of guidelines evaluates current opioid-prescribing recommendations after breast surgery to identify trends in prescribing. Additionally, it compares recommendations on different non-opioid and non-pharmacological adjuncts. METHODS: Electronic databases were searched systematically using terms "breast surgery", "analgesia", "opioid" and "guidelines". The grey literature was used to supplement the search. All articles that provided guidance on opioid prescribing in breast surgery were included. Quality of the guidelines were assessed using the AGREE II tool. Recommendations pertaining to opioid prescribing, analgesic adjuncts and non-pharmacological interventions were summarised and reported with descriptive statistics. RESULT: Eight guidelines pertaining to mastectomies, breast conserving surgery and breast reconstructions were included in this review. Although an opioid-sparing approach was unanimous, there were conflicting recommendations on opioid doses. Opioid requirements were stratified by procedure in 3 guidelines, and by patient risk factors in 2 guidelines. There was significant variability in the recommended multimodal adjuncts. Notably, non-pharmacological interventions such as patient education were infrequently included in guidelines. CONCLUSION: There is a lack of high-quality guidance on opioid prescribing after breast surgery. The optimum approach for personalised opioid prescribing remains unknown. Significant variability between guidelines provide little actionable interventions for prescribers. This could be driven by the paucity in evidence supporting a single efficacious analgesic regimen for patients undergoing breast surgery. Future guidelines should also regularly incorporate non-pharmacological adjuncts to reduce opioid prescribing.

Effect on muscle strength after blood flow restriction resistance exercise in early in-patient rehabilitation of post-chronic obstructive pulmonary disease acute exacerbation, a single blinded, randomized controlled study.

BACKGROUND: Early commencement of rehabilitation might counteract the loss of muscle strength due to a chronic obstructive pulmonary disease acute exacerbation (COPDAE). Blood flow restriction resistance exercise (BFR-RE) using a low intensity of training load has demonstrated muscle strength gain in varieties of clinical populations. This trial aimed at studying the efficacy and acceptability of BFR-RE in patients with post-COPDAE which was not reported before. METHOD: A prospective, assessor blinded, randomized controlled study with 2-week in-patient rehabilitation program with BFR-RE was compared to a matched program with resistance exercise without BFR in patients with post-COPDAE. The primary outcome was the change of muscle strength of knee extensor of dominant leg. The secondary outcomes included changes of hand grip strength (HGS), 6-minute walk test (6MWT) distance, short physical performance battery (SPPB) scores, COPD assessment test (CAT) scores; acceptability and feasibility of BFR-RE; and 1-month unplanned re-admission rate. RESULTS: Forty-Five post-COPDAE patients (mean age = 76 ± 10, mean FEV1%=49% ± 24%) were analyzed. After training, BFR-RE group and control group demonstrated a statistically significant median muscle strength gain of 20 (Interquartile range (IQR) 3 to 38) Newton(N) and 12 (IQR -9 to 30) N respectively. BFR-RE group showed a significant change in SPPB scores, but not in 6MWT distance and HGS after training. Between groups did not have statistically significant different in all primary and secondary outcomes, though with similar acceptability. Drop-out rate due to training-related discomfort in BFR-RE group was 3.7%. CONCLUSION: BFR-RE is feasible and acceptable in patients with post-COPDAE. A 2-week inpatient pulmonary rehabilitation with BFR-RE improved muscle strength of knee extensors, but not a greater extent than the same rehabilitation program with resistance exercise without BFR. Further studies could be considered with a longer training duration and progression of resistance load. [ClinicalTrials.gov Identifier: NCT04448236].

Adapting to children's individual language proficiency: An observational study of preschool teacher talk addressing monolinguals and children learning English as an additional language.

In an increasingly diverse society, young children are likely to speak different first languages that are not the majority language of society. Preschool might be one of the first and few environments where they experience the majority language. The present study investigated how preschool teachers communicate with monolingual English preschoolers and preschoolers learning English as an additional language (EAL). We recorded and transcribed four hours of naturalistic preschool classroom activities and observed whether and how preschool teachers tailored their speech to children of different language proficiency levels and linguistic backgrounds (monolingual English: n = 13; EAL: n = 10), using a suite of tools for analysing quantity and quality of speech. We found that teachers used more diverse vocabulary and more complex syntax with the monolingual children and children who were more proficient in English, showing sensitivity to individual children's language capabilities and adapting their language use accordingly.

A comprehensive molecular phylogeny of the brachyuran crab superfamily Xanthoidea provides novel insights into its systematics and evolutionary history.

Maximum likelihood and Bayesian phylogenies for the brachyuran crab superfamily Xanthoidea were estimated based on three mitochondrial and four nuclear genes to infer phylogenetic relationships and inform taxonomy. Habitat data was then used in conjunction with several diversification rates analyses (BAMM, BiSSE, HiSSE, and FiSSE) to test evolutionary hypotheses regarding the diversification of xanthoid crabs. The phylogenies presented are the most comprehensive to date in terms of global diversity as they include all four constituent families (Xanthidae, Panopeidae, Pseudorhombilidae, and Linnaeoxanthidae) spanning all oceans in which xanthoid crabs occur. Six Xanthoidea families are recognised. Panopeidae and Xanthidae sensu stricto are the two largest family-level clades, which are reciprocally monophyletic. Pseudorhombilidae is nested within and is here treated as a subfamily of Panopeidae. Former subfamilies or tribes of Xanthidae sensu lato are basally positioned clades in Xanthoidea and are here assigned family-level ranks: Garthiellidae, Linnaeoxanthidae, Antrocarcinidae, and Nanocassiopidae. The genera Linnaeoxantho and Melybia were recovered in separate clades with Linnaeoxantho being sister to the family Antrocarcinidae, while Melybia was recovered within the family Panopeidae. The existing subfamily classification of Xanthidae and Panopeidae is drastically restructured with 20 xanthid and four panopeid subfamilies provisionally recognised. Diversification-time analyses inferred the origin of Xanthoidea and Garthiellidae in the Eocene, while the other families originated during the Oligocene. The majority of genus- and species-level diversification took place during the Miocene. Ancestral state reconstruction based on depth of occurrence (shallow vs. deep water) shows some ambiguity for the most recent common ancestor of Xanthoidea and Nanocassiopidae. The most recent common ancestors of Antrocarcinidae and Panopeidae were likely deep-water species, while those of Garthiellidae and Xanthidae were probably shallow-water species. Several shifts in net diversification rates were detected but they were not associated with depth-related habitat transitions.

Functional characterization of a PROTAC directed against BRAF mutant V600E.

The RAF family kinases function in the RAS-ERK pathway to transmit signals from activated RAS to the downstream kinases MEK and ERK. This pathway regulates cell proliferation, differentiation and survival, enabling mutations in RAS and RAF to act as potent drivers of human cancers. Drugs targeting the prevalent oncogenic mutant BRAF(V600E) have shown great efficacy in the clinic, but long-term effectiveness is limited by resistance mechanisms that often exploit the dimerization-dependent process by which RAF kinases are activated. Here, we investigated a proteolysis-targeting chimera (PROTAC) approach to BRAF inhibition. The most effective PROTAC, termed P4B, displayed superior specificity and inhibitory properties relative to non-PROTAC controls in BRAF(V600E) cell lines. In addition, P4B displayed utility in cell lines harboring alternative BRAF mutations that impart resistance to conventional BRAF inhibitors. This work provides a proof of concept for a substitute to conventional chemical inhibition to therapeutically constrain oncogenic BRAF.

Analyzing the impact of CFP1 mutational landscape on epigenetic signaling.

CXXC Zinc finger protein 1 (CFP1) is a multitasking protein playing essential roles during various developmental processes. Its ability to interact with several proteins contribute to several epigenetic events. Here, we review CFP1's functions and its impact on DNA methylation and the post-translational modification of histone proteins such as lysine acetylation and methylation. We will also discuss the potential role of CFP1 in carcinogenesis and the impact of the mutations identified in patients suffering from various cancers.

Gene Flow Increases Phylogenetic Structure and Inflates Cryptic Species Estimations: A Case Study on Widespread Philippine Puddle Frogs (Occidozyga laevis).

In cryptic amphibian complexes, there is a growing trend to equate high levels of genetic structure with hidden cryptic species diversity. Typically, phylogenetic structure and distance-based approaches are used to demonstrate the distinctness of clades and justify the recognition of new cryptic species. However, this approach does not account for gene flow, spatial, and environmental processes that can obfuscate phylogenetic inference and bias species delimitation. As a case study, we sequenced genome-wide exons and introns to evince the processes that underlie the diversification of Philippine Puddle Frogs-a group that is widespread, phenotypically conserved, and exhibits high levels of geographically based genetic structure. We showed that widely adopted tree- and distance-based approaches inferred up to 20 species, compared to genomic analyses that inferred an optimal number of five distinct genetic groups. Using a suite of clustering, admixture, and phylogenetic network analyses, we demonstrate extensive admixture among the five groups and elucidate two specific ways in which gene flow can cause overestimations of species diversity: 1) admixed populations can be inferred as distinct lineages characterized by long branches in phylograms; and 2) admixed lineages can appear to be genetically divergent, even from their parental populations when simple measures of genetic distance are used. We demonstrate that the relationship between mitochondrial and genome-wide nuclear $p$-distances is decoupled in admixed clades, leading to erroneous estimates of genetic distances and, consequently, species diversity. Additionally, genetic distance was also biased by spatial and environmental processes. Overall, we showed that high levels of genetic diversity in Philippine Puddle Frogs predominantly comprise metapopulation lineages that arose through complex patterns of admixture, isolation-by-distance, and isolation-by-environment as opposed to species divergence. Our findings suggest that speciation may not be the major process underlying the high levels of hidden diversity observed in many taxonomic groups and that widely adopted tree- and distance-based methods overestimate species diversity in the presence of gene flow. [Cryptic species; gene flow; introgression; isolation-by-distance; isolation-by-environment; phylogenetic network; species delimitation.].

Repair of multiple simultaneous double-strand breaks causes bursts of genome-wide clustered hypermutation.

A single cancer genome can harbor thousands of clustered mutations. Mutation signature analyses have revealed that the origin of clusters are lesions in long tracts of single-stranded (ss) DNA damaged by apolipoprotein B mRNA editing enzyme, catalytic polypeptide-like (APOBEC) cytidine deaminases, raising questions about molecular mechanisms that generate long ssDNA vulnerable to hypermutation. Here, we show that ssDNA intermediates formed during the repair of gamma-induced bursts of double-strand breaks (DSBs) in the presence of APOBEC3A in yeast lead to multiple APOBEC-induced clusters similar to cancer. We identified three independent pathways enabling cluster formation associated with repairing bursts of DSBs: 5' to 3' bidirectional resection, unidirectional resection, and break-induced replication (BIR). Analysis of millions of mutations in APOBEC-hypermutated cancer genomes revealed that cancer tolerance to formation of hypermutable ssDNA is similar to yeast and that the predominant pattern of clustered mutagenesis is the same as in resection-defective yeast, suggesting that cluster formation in cancers is driven by a BIR-like mechanism. The phenomenon of genome-wide burst of clustered mutagenesis revealed by our study can play an important role in generating somatic hypermutation in cancers as well as in noncancerous cells.

Evaluation of a SARS-CoV-2 Surrogate Virus Neutralization Test for Detection of Antibody in Human, Canine, Cat, and Hamster Sera.

Surrogate neutralization assays for severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) that can be done without biosafety level 3 containment and in multiple species are desirable. We evaluate a recently developed surrogate virus neutralization test (sVNT) in comparison to 90% plaque reduction neutralization tests (PRNT90) in human, canine, cat, and hamster sera. With PRNT90 as the reference, sVNT had sensitivity of 98.9% and specificity of 98.8%. Using a panel of immune sera corresponding to other coronaviruses, we confirm the lack of cross-reactivity to other coronaviruses in SARS-CoV-2 sVNT and PRNT90, except for cross-reactivity to SARS-CoV-1 in sVNT.

Comparison of SARS-CoV-2 indirect and direct RT-qPCR detection methods.

BACKGROUND: Sensitive, rapid, and accessible diagnostics continue to be critical to track the COVID-19 pandemic caused by the SARS-CoV-2 virus. RT-qPCR is the gold standard test, and comparison of methodologies and reagents, utilizing patient samples, is important to establish reliable diagnostic pipelines. METHODS: Here, we assessed indirect methods that require RNA extraction with direct RT-qPCR on patient samples. Four different RNA extraction kits (Qiagen, Invitrogen, BGI and Norgen Biotek) were compared. For detection, we assessed two recently developed Taqman-based modules (BGI and Norgen Biotek), a SYBR green-based approach (NEB Luna Universal One-Step Kit) with published and newly-developed primers, and clinical results (Seegene STARMag RNA extraction system and Allplex 2019-nCoV RT-qPCR assay). We also tested and optimized direct, extraction-free detection using these RT-qPCR systems and performed a cost analysis of the different methods evaluated here. RESULTS: Most RNA isolation procedures performed similarly, and while all RT-qPCR modules effectively detected purified viral RNA, the BGI system provided overall superior performance (lower detection limit, lower Ct values and higher sensitivity), generating comparable results to original clinical diagnostic data, and identifying samples ranging from 65 copies to 2.1 × 105 copies of viral genome/µl. However, the BGI detection system is more expensive than other options tested here. With direct RT-qPCR, simply adding an RNase inhibitor greatly improved detection, without the need for any other treatments (e.g. lysis buffers or boiling). The best direct methods detected ~ 10 fold less virus than indirect methods, but this simplified approach reduced sample handling, as well as assay time and cost. CONCLUSIONS: With extracted RNA, the BGI RT-qPCR detection system exhibited superior performance over the Norgen system, matching initial clinical diagnosis with the Seegene Allplex assay. The BGI system was also suitable for direct, extraction-free analysis, providing 78.4% sensitivity. The Norgen system, however, still accurately detected samples with a clinical Ct < 33 from extracted RNA, provided significant cost savings, and was superior to SYBR green assays that exhibited reduced specificity.

Role of caregiver factors in outpatient medical follow-up post-stroke: observational study in Singapore.

BACKGROUND: Outpatient medical follow-up post-stroke is not only crucial for secondary prevention but is also associated with a reduced risk of rehospitalization. However, being voluntary and non-urgent, it is potentially determined by both healthcare needs and the socio-demographic context of stroke survivor-caregiver dyads. Therefore, we aimed to examine the role of caregiver factors in outpatient medical follow-up (primary care (PC) and specialist outpatient care (SOC)) post-stroke. METHOD: Stroke survivors and caregivers from the Singapore Stroke Study, a prospective, yearlong, observational study, contributed to the study sample. Participants were interviewed 3-monthly for data collection. Counts of PC and SOC visits were extracted from the National Claims Database. Poisson modelling was used to explore the association of caregiver (and patient) factors with PC/SOC visits over 0-3 months (early) and 4-12 months (late) post-stroke. RESULTS: For the current analysis, 256 stroke survivors and caregivers were included. While caregiver-reported memory problems of a stroke survivor (IRR: 0.954; 95% CI: 0.919, 0.990) and caregiver burden (IRR: 0.976; 95% CI: 0.959, 0.993) were significantly associated with lower early post-stroke PC visits, co-residing caregiver (IRR: 1.576; 95% CI: 1.040, 2.389) and negative care management strategies (IRR: 1.033; 95% CI: 1.005, 1.061) were significantly associated with higher late post-stroke SOC visits. CONCLUSION: We demonstrated that the association of caregiver factors with outpatient medical follow-up varied by the type of service (i.e., PC versus SOC) and temporally. Our results support family-centred care provision by family physicians viewing caregivers not only as facilitators of care in the community but also as active members of the care team and as clients requiring care and regular assessments.

The relationship between PD-L1 expression and tumor driver gene mutations and survival in non small cell lung carcinoma.

To explore the relationship between PD-L1 expression and gene mutations and survival. PD L1, ALK and MET protein expression were detected by immunohistochemistry, and EGFR gene mutation by RT-PCR in 209 cases of NSCLC. The correlations between PD-L1 expression and gene mutations, clinicopathological features and survival was analyzed. PD-L1 was expressed in 99/209 cases (47.4%) of NSCLC, including score 1 (≥ 1% to < 5%) 23 cases (11%), score 2 (≥ 5% to 50%) 40 cases (19.1%). There were 89 cases (42.6%) of EGFR mutation, 12 (5.7%) of ALK and 90 (43.1%) of MET protein positive. PD-L1 positive expression occurred more frequently in men and non-adenocarcinoma, and was negatively correlated with EGFR mutation and histological differentiation of NSCLC. PD-L1 expression was concordant in primary and metastatic cancers. There was no any effect of PD-L1 expression on overall survival of patients with NSCLC. These results suggested that PD-L1 expression is not an independent risk factor for survival of patients with NSCLC. Because of mutual complementarities of PD-L1 expression and EGFR mutation in NSCLC, both should be simultaneously detected for the patients to achieve eligible treatments.