Examining the approaches used to assess decision-making capacity in healthcare practice.

AIM: To examine the approaches that are being used in New Zealand when conducting decision-making capacity (DMC) assessments among the healthcare professionals that commonly conduct DMC assessments and those that are involved in, but do not conduct, the assessments. METHOD: An online quantitative survey was conducted, lasting 10 minutes, including a mix of closed- and open-ended questions. The survey garnered responses from a total of n=78 participants. RESULTS: Bedside cognitive tests were found to be the most commonly reported tool used to assess DMC among those conducting and those contributing to DMC assessments. Nearly a third (31.9%) of participants conducting DMC assessments used a structured clinical interview as one of their most common approaches while 27.5% of this same group reported not being aware of this approach. It was reported by both those conducting and those contributing to DMC assessments that the current standards lack quality and consistency, with partial capacity being poorly understood and identified, and supported decision making often being overlooked for substitute decision making. CONCLUSIONS: Current approaches to DMC assessment lack standardisation and consistency, with assessment approaches being widely varied. This article serves as a call for the development of and adherence to nationally recognised standards for DMC assessments.

Epidemiology of Faecal Incontinence for People with Dementia Living in the Community in New Zealand: A Retrospective Cohort Study Using interRAI Home Care Assessment Data.

INTRODUCTION: Globally there are few studies but wide variation in epidemiology of faecal incontinence (FI) for people living with dementia in the community. Our objectives are to identify 1-year period prevalence, 5-year incidence, and risks for faecal incontinence (FI) for people living with dementia. METHODS: A retrospective cohort study comprising International Residential Assessment Instrument Home Care version (interRAI-HC) assessments in a 5-year period in New Zealand (N=109964). For prevalence analysis, we selected a dementia cohort for a 1-year period from 1 Aug 2020 to 31 July 2021 (n=7775). For the incidence analysis, participants in the dementia cohort were followed up from the day of the first dementia diagnosis during the period 1 Aug 2016 and 31 July 2021. Dementia was identified by combining diagnosis of "Alzheimer's disease" and "Dementia other than Alzheimer's disease". Participants were coded with faecal incontinence if they were continent with a stoma, seldom incontinent, occasionally incontinent, often incontinent and incontinent. RESULTS: 1 year period (1 Aug 2020-31 July 2021) prevalence of FI was 26.7% (2082/7775) of PLWD. 5-year incident FI rate was 19.0 per 100 person years for PLWD and 12.3 per 100 person years for people without dementia. Controlling for risk factors for FI in both groups the hazard ratio for FI was 1.7 for PLWD. CONCLUSION: FI affects a significant proportion of people with dementia in NZ. interRAI-HC data could facilitate global epidemiological studies to estimate service or intervention need for people with dementia to redress or manage FI.

Identifying Frailty in Older Adults Receiving Home Care Assessment Using Machine Learning: Longitudinal Observational Study on the Role of Classifier, Feature Selection, and Sample Size.

BACKGROUND: Machine learning techniques are starting to be used in various health care data sets to identify frail persons who may benefit from interventions. However, evidence about the performance of machine learning techniques compared to conventional regression is mixed. It is also unclear what methodological and database factors are associated with performance. OBJECTIVE: This study aimed to compare the mortality prediction accuracy of various machine learning classifiers for identifying frail older adults in different scenarios. METHODS: We used deidentified data collected from older adults (65 years of age and older) assessed with interRAI-Home Care instrument in New Zealand between January 1, 2012, and December 31, 2016. A total of 138 interRAI assessment items were used to predict 6-month and 12-month mortality, using 3 machine learning classifiers (random forest [RF], extreme gradient boosting [XGBoost], and multilayer perceptron [MLP]) and regularized logistic regression. We conducted a simulation study comparing the performance of machine learning models with logistic regression and interRAI Home Care Frailty Scale and examined the effects of sample sizes, the number of features, and train-test split ratios. RESULTS: A total of 95,042 older adults (median age 82.66 years, IQR 77.92-88.76; n=37,462, 39.42% male) receiving home care were analyzed. The average area under the curve (AUC) and sensitivities of 6-month mortality prediction showed that machine learning classifiers did not outperform regularized logistic regressions. In terms of AUC, regularized logistic regression had better performance than XGBoost, MLP, and RF when the number of features was ≤80 and the sample size ≤16,000; MLP outperformed regularized logistic regression in terms of sensitivities when the number of features was ≥40 and the sample size ≥4000. Conversely, RF and XGBoost demonstrated higher specificities than regularized logistic regression in all scenarios. CONCLUSIONS: The study revealed that machine learning models exhibited significant variation in prediction performance when evaluated using different metrics. Regularized logistic regression was an effective model for identifying frail older adults receiving home care, as indicated by the AUC, particularly when the number of features and sample sizes were not excessively large. Conversely, MLP displayed superior sensitivity, while RF exhibited superior specificity when the number of features and sample sizes were large.

Sorted-Cell Sequencing on HCC Specimens Reveals EPS8L3 as a Key Player in CD24/CD13/EpCAM-Triple Positive, Stemness-Related HCC Cells.

BACKGROUND & AIMS: Hepatocellular carcinoma (HCC) is a heterogeneous cancer with varying levels of liver tumor initiating or cancer stem cells in the tumors. We aimed to investigate the expression of different liver cancer stem cell (LCSC) markers in human HCCs and identify their regulatory mechanisms in stemness-related cells. METHODS: We used an unbiased, single-marker sorting approach by flow cytometry, fluorescence-activated cell sorting, and transcriptomic analyses on HCC patients' resected specimens. Knockdown approach was used, and relevant functional assays were conducted on the identified targets of interest. RESULTS: Flow cytometry on a total of 60 HCC resected specimens showed significant heterogeneity in the expression of LCSC markers, with CD24, CD13, and EpCAM mainly contributing to this heterogeneity. Concomitant expression of CD24, CD13, and EpCAM was detected in 32 HCC samples, and this was associated with advanced tumor stages. Transcriptomic sequencing on the HCC cells sorted for these individual markers identified epidermal growth factor receptor kinase substrate 8-like protein 3 (EPS8L3) as a common gene associated with the 3 markers and was functionally validated in HCC cells. Knocking down EPS8L3 suppressed the expression of all 3 markers. To search for the upstream regulation of EPS8L3, we found SP1 bound to EPS8L3 promoter to drive EPS8L3 expression. Furthermore, using Akt inhibitor MK2206, we showed that Akt signaling-driven SP1 drove the expression of the 3 LCSC markers. CONCLUSIONS: Our findings suggest that Akt signaling-driven SP1 promotes EPS8L3 expression, which is critical in maintaining the downstream expression of CD24, CD13, and EpCAM. The findings provide insight into potential LCSC-targeting therapeutic strategies.

Exploring training, involvement and confidence: a study of healthcare professionals in decision-making capacity assessments.

AIM: To explore the training, involvement and confidence of healthcare professionals involved in decision-making capacity (DMC) assessments, and to compare any differences between those conducting and those involved in, but not conducting DMC assessments. METHOD: A 10-minute anonymous, online survey was conducted with both closed and open questions. A total of 78 participants completed the survey. RESULTS: Training was lacking in quantity and adequacy. Only 14.1% received formal training during and post their qualification and only 38.5% reported the right amount of training. Just over 55% reported having the right amount of involvement, with 18% having too much and 27% having not enough involvement. A significantly higher response was given for having too much involvement by those conducting DMC assessments (p=0.006), while those not conducting felt they do not have enough involvement (p<0.001). Only 25.6% (n=20) were very confident in being able to explain DMC to a patient. CONCLUSIONS: Healthcare professionals working in this area urgently require support in the form of formal training and defined roles. Given what can be at stake for an individual undergoing a DMC assessment, it is imperative that improvements are made to upskill the workforce and utilise expertise of all healthcare professionals.

Using interRAI Assessment for Research: Developing a National Research Agenda in Aotearoa New Zealand.

interRAI provides a suite of standardized, validated instruments used to assess health and psychosocial well-being, and to inform person-centered care planning. Data obtained from these standardized tools can also be used at a population level for research and to inform policy, and interRAI is currently used in more than 40 countries globally. We present a brief overview of the use of interRAI internationally within research and policy settings, and then introduce how interRAI is used within the universal public health system in Aotearoa New Zealand (NZ), including considerations relating to Maori, the Indigenous people of NZ. In NZ, improvement in interRAI data utilization for research purposes was called for from aged care, health providers, and researchers, to better use these data for quality improvement and health advancement for New Zealanders. A national research network has been established, providing a medium for researchers to form relationships and collaborate on interRAI research with a goal of translating routinely collected interRAI data to improve clinical care, patient experience, service development, and quality improvement. In 2023, the network members met (hybrid: in-person and online) and identified research priorities. These were collated and developed into a national interRAI research agenda by the NZ interRAI Research Network Working Group. Research priorities included reviewing the interRAI assessment processes, improving methods for data linkage to national data sets, exploring how Indigenous Data Sovereignty can be upheld, as well as a variety of clinically focused research topics. Implications for Practice, Policy, and Research: This appears to be the first time national interRAI research priorities have been formally identified. Priorities identified have the potential to inform quality and clinical improvement activities and are likely of international relevance. The methodology described to cocreate the research priorities will also be of wider significance for those looking to do so in other countries.

Penetration of Calcium Silicate and Epoxy Resin Sealers Into the Lateral Canals.

OBJECTIVES: The aim of this research was to compare the penetration ability of calcium silicate-based sealers (iRoot SP and TotalFill BC HiFlow) and an epoxy resin-based sealer (AH Plus) into the lateral canals using the single-cone (SC) or continuous wave compaction (CW) obturation techniques. METHODS: Ninety single-rooted human teeth received canal instrumentation and diaphanisation before artificial lateral canals were created at 3 different root levels. The specimens were randomly allocated into 6 groups based on the combination of obturation technique and sealer used. Radiographic and photographic measurements evaluated the percentage of sealer penetration. Statistical analysis was performed to compare the penetration ability amongst different types of sealers, obturation methods, and root levels. RESULTS: Although TotalFill BC HiFlow sealer combined with the CW technique demonstrated greater sealer penetration at the coronal level (P < .05), the overall penetration ability of iRoot SP, TotalFill BC HiFlow, and AH Plus was not significantly different. The deepest sealer penetration was observed at the apical root level. CONCLUSIONS: All sealers showed similar penetration abilities into the lateral canals using the SC or CW techniques in vitro. Calcium silicate-based sealers have comparable penetration ability into lateral canals compared to epoxy resin-based sealers using SC or CW obturation techniques.

Patterns of opioid use in New Zealand older adults, 2007-2018.

OBJECTIVES: Opioid use has increased globally, dramatically increasing opioid overdose, dependence, abuse and mortality. Limited research is available on opioid use patterns in older adults in New Zealand and internationally. This study aims to address this gap by determining the incidence and prevalence of opioid use among older adults (age ≥65 years) in New Zealand from 2007 to 2018. METHODS: This was a population-based retrospective cohort study conducted using New Zealand national administrative healthcare databases. The annual opioid use incidence (2008-2018) and prevalence (2007-2018) in older adults were determined and stratified by sex, age, and opioid type and strength. We used descriptive statistics to summarise the patterns of opioid dispensing. Data analysis was conducted using MS Excel, and data linking was performed using SQL software. RESULTS: A total of 820,349 older adults were initiated on opioids during the study period. The overall incidence of opioid use in older adults showed a steady increase from 2008 to 2015; similarly, the prevalence steadily increased from 2007 to 2015, and thereafter, both rates fluctuated. A slight decrease in both prevalence and incidence rates was observed in 2018. Codeine and tramadol were the most commonly dispensed opioids during the study period. Females had a higher incidence and prevalence of all opioids than males. CONCLUSIONS: The incidence and prevalence of opioid dispensing increased in New Zealand older adults over time. Monitoring the trends of opioid use in older adults is critical to enable clinicians and policymakers to deliver early interventions to prevent future opioid-related adverse events.

Reciprocal interactions between malignant cells and macrophages enhance cancer stemness and M2 polarization in HBV-associated hepatocellular carcinoma.

Background: The tumor microenvironment of cancers has emerged as a crucial component in regulating cancer stemness and plays a pivotal role in cell-cell communication. However, the specific mechanisms underlying these phenomena remain poorly understood. Methods: We performed the single-cell RNA sequencing (scRNA-seq) on nine HBV-associated hepatocellular carcinoma (HCC) patients. The heterogeneity of the malignant cells in pathway functions, transcription factors (TFs) regulation, overall survival, stemness, as well as ligand-receptor-based intercellular communication with macrophages were characterized. The aggressive and stemness feature for the target tumor subclone was validated by the conduction of in vitro assays including sphere formation, proliferation, Annexin V apoptosis, flow cytometry, siRNA library screening assays, and multiple in vivo preclinical mouse models including mouse hepatoma cell and human HCC cell xenograft models with subcutaneous or orthotopic injection. Results: Our analysis yielded a comprehensive atlas of 31,664 cells, revealing a diverse array of malignant cell subpopulations. Notably, we identified a stemness-related subclone of HCC cells with concurrent upregulation of CD24, CD47, and ICAM1 expression that correlated with poorer overall survival. Functional characterization both in vitro and in vivo validated S100A11 as one of the top downstream mediators for tumor initiation and stemness maintenance of this subclone. Further investigation of cell-cell communication within the tumor microenvironment revealed a propensity for bi-directional crosstalk between this stemness-related subclone and tumor-associated macrophages (TAMs). Co-culture study showed that this interaction resulted in the maintenance of the expression of cancer stem cell markers and driving M2-like TAM polarization towards a pro-tumorigenic niche. We also consolidated an inverse relationship between the proportions of TAMs and tumor-infiltrating T cells. Conclusions: Our study highlighted the critical role of stemness-related cancer cell populations in driving an immunosuppressive tumor microenvironment and identified the S100A11 gene as a key mediator for stemness maintenance in HCC. Moreover, our study provides support that the maintenance of cancer stemness is more attributed to M2 polarization than the recruitment of the TAMs.

LANCL1, a cell surface protein, promotes liver tumor initiation through FAM49B-Rac1 axis to suppress oxidative stress.

BACKGROUND AND AIMS: HCC is an aggressive cancer with a poor clinical outcome. Understanding the mechanisms that drive tumor initiation is important for improving treatment strategy. This study aimed to identify functional cell membrane proteins that promote HCC tumor initiation. APPROACH AND RESULTS: Tailor-made siRNA library screening was performed for all membrane protein-encoding genes that are upregulated in human HCC (n = 134), with sphere formation as a surrogate readout for tumor initiation. Upon confirmation of membranous localization by immunofluorescence and tumor initiation ability by limiting dilution assay in vivo, LanC-like protein-1 (LANCL1) was selected for further characterization. LANCL1 suppressed intracellular reactive oxygen species (ROS) and promoted tumorigenicity both in vitro and in vivo. Mechanistically, with mass spectrometry, FAM49B was identified as a downstream binding partner of LANCL1. LANCL1 stabilized FAM49B by blocking the interaction of FAM49B with the specific E3 ubiquitin ligase TRIM21, thus protecting FAM49B from ubiquitin-proteasome degradation. The LANCL1-FAM49B axis suppressed the Rac1-NADPH oxidase-driven ROS production, but this suppression of ROS was independent of the glutathione transferase function of LANCL1. Clinically, HCCs with high co-expression of LANCL1 and FAM49B were associated with more advanced tumor stage, poorer overall survival, and disease-free survival. In addition, anti-LANCL1 antibodies targeting the extracellular N-terminal domain were able to suppress the self-renewal ability, as demonstrated by the sphere formation ability of HCC cells. CONCLUSIONS: Our data showed that LANCL1 is a cell surface protein and a key contributor to HCC initiation. Targeting the LANCL1-FAM49B-Rac1-NADPH oxidase-ROS signaling axis may be a promising therapeutic strategy for HCC.

Falls Risk in Long-Term Care Residents With Cognitive Impairment: Effects of COVID-19 Pandemic.

OBJECTIVES: The aim of this study was to investigate the impact of the COVID-19 pandemic on falls rates in long-term care residents with cognitive impairment. DESIGN: An observational study using routinely collected national interRAI data. SETTING AND PARTICIPANTS: Participants were from long-term care residents (age ≥60 years) who received an interRAI Long Term Care Facility assessment anywhere in New Zealand between August 17, 2018, and August 16, 2022. METHODS: The primary outcome was "At least 1 fall in the last 30 days." Based on the Cognitive Performance Scale (CPS), cognitive impairment was categorized into 3 levels: intact or borderline intact (0-1), mild to moderate impairment (2-3), and moderately to very severe impairment (4-6). The COVID-19 pandemic was divided into 3 periods (First wave: March 21, 2020, to June 8, 2020; Varying level of community outbreaks: June 9, 2020 to August 16, 2021; and Delta-Omicron wave: August 17, 2021, to August 16, 2021) and compared to a pre-COVID-19 period (August 17, 2018, to March 20, 2020). Cox regression modeling was used to study falls and interactions between CPS and COVID-19 pandemic periods, along with other established falls risk factors in the literature. RESULTS: A total of 282,518 interRAI-LTCF assessments from 75,132 unique residents were included. Interactions between CPS and COVID-19 pandemic periods found that cognitive impairment was associated with a higher hazard ratio (ranged from 1.22 to 1.37) in each of the 3 COVID-19 pandemic periods. We also found unstable health, unsteady gait, wandering, and moderate to severe ADL dependency were the strongest risk factors for falls. CONCLUSIONS AND IMPLICATIONS: Cognitively impaired long-term care residents had an increased risk for falls during the COVID-19 pandemic. This risk was influenced by several factors. In future pandemic or infection control related isolation, residents who are most at risk can be identified for targeted falls prevention programs.

Reliability and reproducibility of CBCT assessment of mandibular changes before and after treatment for Class III growing patients - an easy and quick way for evaluation.

The objective of this study was to evaluate intraobserver reliability and inter-observer reproducibility of a 3-dimensional (3D) assessment method for mandibular changes of growing patients after orthodontic treatment for Class III malocclusion.Methods Cone-beam computed tomography (CBCT) scans were performed before and after orthodontic treatment for 27 patients. During the scan, the patient was positioned such that his/her mandibular plane was parallel to floor. Three observers independently worked on the DICOM data, reconstructed the pre- and post-treatment 3D models in software, selected the stable anatomical structures (basal bone area from the lingual surface of the symphysis to the distal aspect of the first molars) to guide the automated superimposition process. Then, each observer registered 14 anatomical landmarks on the virtual models, for three times after suitable interval, to generate 3 sets of coordinates; the mean was taken as the coordinates for that particular landmark. The intraobserver reliability and inter-observer reproducibility of the method were analyzed.Results The ICCs was > 0.90 for 25 (92.6%) of the intraobserver assessments. The precision of the measurement method was < 0.3 mm in 24 (88.9%) cases. The interobserver reproducibility errors were < 0.3 mm in 21 of the 27 cases.Conclusions The intraobserver reliability and inter-observer reproducibility of 3D assessment of mandibular changes using the virtual models were excellent.

Tissue Stabilization, Bacterial Adhesion, and Stem Cell Viability in Trans-cinnamaldehyde-conditioned Dentin.

INTRODUCTION: This laboratory study aimed to evaluate the effect of trans-cinnamaldehyde (TC) conditioning on dentin tissue stabilization, bacterial adhesion, and stem cell toxicity. METHODS: Dentin beams (n = 204) from extracted human molars were demineralized in phosphoric acid and treated with TC (2.5, 5, and 7.5%), 50% ethanol-water mixture (vehicle control) or 2.5% glutaraldehyde (GA) (positive control) for 30 minutes. Demineralized but untreated specimens served as the negative control. After treatment, collagen crosslinking was characterized by measuring the elastic modulus (Er) and hardness (n = 5). Biodegradation resistance was examined by determining the loss of dry mass (n = 8), hydroxyproline release (n = 4) and scanning electron microscopy (n = 2), after exposure to bacterial collagenase. Inhibition of bacterial adhesion was investigated by colony counting assay (n = 12) and scanning electron microscopy (n = 2). Viability of stem cells of the apical papilla on TC-conditioned dentin was determined using the Cell Counting Kit-8 assay (n = 8). Data were statistically analyzed using one-way analysis of variance (ANOVA) test followed by Dunnett's multiple comparisons at a significance level of 5%. RESULTS: TC-conditioned dentin showed a concentration-dependent increase in Er and hardness. The Er and hardness of 5% and 7.5% TC-conditioned dentin were significantly greater than that of the negative control and vehicle control groups (P < .05). There was no significant difference in the biodegradation resistance between GA and 5% TC-conditioned dentin (P > .05). TC-conditioned dentin showed a well-preserved collagen fibril network with clear cross-banding, comparable to GA-conditioned dentin. All concentrations of TC inhibited bacterial adhesion on dentin, significantly greater than the negative control (P < .05). There was no reduction in viability of stem cells of the apical papilla viability on TC-conditioned dentin compared to the negative control (P > .05). CONCLUSIONS: TC conditioning stabilized the dentin and protected it from enzymatic degradation. TC prevented bacterial adhesion on the dentin but maintained stem cell viability.

Speech and language processing with deep learning for dementia diagnosis: A systematic review.

Dementia is a progressive neurodegenerative disease that burdens the person living with the disease, their families, and medical and social services. Timely diagnosis of dementia could be followed by introducing interventions that may slow down its progression or reduce its burdens. However, the diagnostic process of dementia is often complex and resource intensive. Access to diagnostic services is also an issue in low and middle-income countries. The abundance and easy accessibility of speech and language data have created new possibilities for utilizing Deep Learning (DL) technologies to be part of the dementia diagnostic process. This systematic review included studies published between 2012-2022 that utilized such technologies to aid in diagnosing dementia. We identified 72 studies using the PRISMA 2020 protocol, extracted and analyzed data from these studies and reported the related DL technologies. We found these technologies effectively differentiated between healthy individuals and those with a dementia diagnosis, highlighting their potential in the diagnosis of dementia. This systematic review provides insights into the contributions of DL-based speech and language techniques to support the dementia diagnostic process. It also offers an understanding of the advancements made in this field thus far and highlights some challenges that still need to be addressed.

Predictors of persistent opioid use in non-cancer older adults: a retrospective cohort study.

BACKGROUND: Long-term opioid use and associated adverse outcomes have increased dramatically in recent years. Limited research is available on long-term opioid use in older adults. OBJECTIVE: We aimed to determine the incidence and predictors of long-term or persistent opioid use (POU) amongst opioid-naïve older adults without a cancer diagnosis. METHODS: This was a retrospective cohort study using five national administrative healthcare databases in New Zealand. We included all opioid-naïve older adults (≥65 years) who were initiated on opioid therapy between January 2013 and June 2018. The outcome of interest was POU, defined as having continuously filled ≥1 opioid prescription within 91-180 days after the index opioid prescription. Multivariable logistic regression was used to examine the predictors of POU. RESULTS: The final sample included 268,857 opioid-naïve older adults; of these, 5,849(2.2%) developed POU. Several predictors of POU were identified. The use of fentanyl (adjusted odds ratio (AOR) = 3.61; 95% confidence interval (CI) 2.63-4.95), slow-release opioids (AOR = 3.02; 95%CI 2.78-3.29), strong opioids (AOR = 2.03; 95%CI 1.55-2.65), Charlson Comorbidity Score ≥ 3 (AOR = 2.09; 95% CI 1.78-2.46), history of substance abuse (AOR = 1.52; 95%CI 1.35-1.72), living in most socioeconomically deprived areas (AOR = 1.40; 95%CI 1.27-1.54), and anti-epileptics (AOR = 2.07; 95%CI 1.89-2.26), non-opioid analgesics (AOR = 2.05; 95%CI 1.89-2.21), antipsychotics (AOR = 1.96; 95%CI 1.78-2.17) or antidepressants (AOR = 1.50; 95%CI 1.41-1.59) medication use were the strongest predictors of POU. CONCLUSION: A significant proportion of patients developed POU, and several factors were associated with POU. The findings will enable healthcare providers and policymakers to target early interventions to prevent POU and related adverse events.

Exploring the impact of e-learning modules and webinars on health professionals' understanding of the End of Life Choice Act 2019: a secondary analysis of Manatu Hauora - Ministry of Health workforce survey.

AIM: To explore the importance of health workforce training, particularly in newly regulated healthcare practices such as assisted dying (AD). This study aims to analyse the socio-demographic factors associated with health professionals' completion of the e-learning module and attendance at the two webinars provided by the New Zealand Ministry of Health - Manatu Hauora (MH) and whether completion of the e-learning module and webinars supported health professionals' understanding of the End of Life Choices Act 2019. METHOD: Secondary analysis of the MH workforce surveys conducted in July 2021. RESULTS: The study findings indicate that health professionals who are older, of Pakeha/European ethnicity and work in hospice settings are more likely to complete the e-learning module, while females are more likely to attend webinars. CONCLUSION: Despite low completion and attendance rates, the study highlights the positive association between training and health professionals' overall understanding of the Act. These results emphasise the need for enhancing training programmes to increase health professionals' knowledge and competence with AD. Furthermore, the research proposes focussing on healthcare practitioners in the early stages of their careers and not directly engaged in offering AD services, as well as Maori and Pasifika health practitioners.

Intracellular bacterial eradication using a novel peptide in vitro.

AIM: To determine the effect of a novel antimicrobial peptide (AMP; OP145) and cell-penetrating peptide (Octa-arginine/R8) conjugate on the killing of intracellular Enterococcus faecalis, compared to OP145 and an antibiotic combination recommended for regenerative endodontic procedures. METHODOLOGY: The biocompatible concentrations of OP145 and OP145-R8 were determined by assessing their cytotoxicity against human macrophages and red blood cells. Spatiotemporal internalization of the peptides into macrophages was investigated qualitatively and quantitatively by confocal laser scanning microscopy and flow cytometry respectively. Killing of extracellular and intracellular E. faecalis OG1RF by the peptides was determined by counting the colony-forming units (CFU). Intracellular antibacterial activity of the peptides was compared to a double antibiotic combination. Confocal microscopy was used to confirm the intracellular bacterial eradication. Significant differences between the different test groups were analysed using one-way analysis of variance. p < .05 was considered to be statistically significant. RESULTS: Peptides at a concentration of 7.5 µmol/L were chosen for subsequent experiments based on the results of the alamarBlue™ cell viability assay and haemolytic assay. OP145-R8 selectively internalized into lysosomal compartments and the cytosol of macrophages. Conjugation with R8 improved the internalization of OP145 into macrophages in a temporal manner (70.53% at 1 h to 77.13% at 2 h), while no temporal increase was observed for OP145 alone (60.53% at 1 h with no increase at 2 h). OP145-R8 demonstrated significantly greater extracellular and intracellular antibacterial activity compared to OP145 at all investigated time-points and concentrations (p < .05). OP145-R8 at 7.5 µmol/L eradicated intracellular E. faecalis after 2 h (3.5 log reduction compared to the control; p < .05), while the antibiotics could not reduce more than 0.5 log CFU compared to the control (p > .05). Confocal microscopy showed complete absence of E. faecalis within the OP145-R8 treated macrophages. CONCLUSIONS: The results of this study demonstrated that the conjugation of an AMP OP145 to a cell-penetrating peptide R8 eradicated extracellular and intracellular E. faecalis OG1RF without toxic effects on the host cells.