Smoking cessation problem-based learning: Virtual experience

Background and Objectives: Problem-based learning (PBL) is a student-centered teaching and learning methodology where students collaboratively address specific issues. Tobacco use is a major health issue globally. Health professions and students need to have knowledge and skills to facilitate smoking cessation. The objective of this study is to assess feasibility of PBL during a virtual attachment involving institutions from Malaysia and the USA. Methods: A 4-week smoking cessation virtual attachment was conducted for three third-year University of Pittsburgh, USA pharmacy students. Malaysian smoking cessation experts designed and facilitated a PBL smoking cessation module. It was split into two 2-hour sessions with 3 triggers; Trigger 1: ‘Chief Presentation’, Trigger 2: ‘History & Motivational Interview’, and Trigger 3: ‘Brief 5A’s Intervention’. Students received Trigger 1 a day earlier and discussed amongst themselves. In session 1, Triggers 1-3 were given sequentially and discussed after completing all tasks from each trigger. In session 2 one-week later, facilitators gave formative assessment and students provided reflection regarding the PBL session. Upon completing the four-week virtual attachment, students provided feedback and facilitators graded the students. Result and Discussion: A comprehensive and interactive PBL session was successfully conducted virtually. Based on the clinical practice guidelines of both countries, there were differences in terms of availability and use of cessation medications, but the general principles of smoking cessation consultation and interventions were similar. Students were able to discuss the case openly, putting forth ideas and questions in both sessions. All students provided positive feedbacks regarding the PBL. Conclusions: With the extensive development of online platforms connecting the world over, student virtual attachment and mobility programmes can be easily conducted with minimal cost. A suitable module embedding PBL can be designed and conducted to best suit the online platform and the intended students

Separator fluid volume requirements in multi-infusion settings

INTRODUCTION. Intravenous (IV) therapy is a widely used method for the administration of medication in hospitals worldwide. ICU and surgical patients in particular often require multiple IV catheters due to incompatibility of certain drugs and the high complexity of medical therapy. This increases discomfort by painful invasive procedures, the risk of infections and costs of medication and disposable considerably. When different drugs are administered through the same lumen, it is common ICU practice to flush with a neutral fluid between the administration of two incompatible drugs in order to optimally use infusion lumens. An important constraint for delivering multiple incompatible drugs is the volume of separator fluid that is sufficient to safely separate them. OBJECTIVES. In this pilot study we investigated whether the choice of separator fluid, solvent, or administration rate affects the separator volume required in a typical ICU infusion setting. METHODS. A standard ICU IV line (2m, 2ml, 1mm internal diameter) was filled with methylene blue (40 mg/l) solution and flushed using an infusion pump with separator fluid. Independent variables were solvent for methylene blue (NaCl 0.9% vs. glucose 5%), separator fluid (NaCl 0.9% vs. glucose 5%), and administration rate (50, 100, or 200 ml/h). Samples were collected using a fraction collector until <2% of the original drug concentration remained and were analyzed using spectrophotometry. RESULTS. We did not find a significant effect of administration rate on separator fluid volume. However, NaCl/G5% (solvent/separator fluid) required significantly less separator fluid than NaCl/NaCl (3.6 ± 0.1 ml vs. 3.9 ± 0.1 ml, p <0.05). Also, G5%/G5% required significantly less separator fluid than NaCl/NaCl (3.6 ± 0.1 ml vs. 3.9 ± 0.1 ml, p <0.05). The significant decrease in required flushing volume might be due to differences in the viscosity of the solutions. However, mean differences were small and were most likely caused by human interactions with the fluid collection setup. The average required flushing volume is 3.7 ml. CONCLUSIONS. The choice of separator fluid, solvent or administration rate had no impact on the required flushing volume in the experiment. Future research should take IV line length, diameter, volume and also drug solution volumes into account in order to provide a full account of variables affecting the required separator fluid volume

Defining the human mucosal CD4+ T cell and Mononuclear Phagocyte landscape by high parameter technologies

Tissue resident mononuclear phagocytes (MNP) are among the first immune cells that sexually transmitted pathogens encounter during transmission. These antigen presenting cells rapidly endocytose pathogens and then interact with CD4+ T cells to initiate adaptive immunity. In the case of HIV, the virus can be transferred from MNP to infect CD4+ T cells which are the primary target cell in which HIV undergoes replication. These MNP:CD4+ T cell interaction were originally believed to occur in lymph nodes but there is now an increasing body of evidence that this occurs within tissue at the site of transmission. There has been recent interest in defining the phenotype and the role of CD4+ TRMs in diseases. In HIV, CD4+ TRM have been shown to be a major target for productive and latent infection in the cervix. However, there are limited investigations into the CD4+ T cell landscape in other genital tissues. In this thesis, we optimised the enzymatic digestion process to maintain the cells in their physiological status. We have designed, optimised and implemented a 24-color flow cytometry panel to characterise T cells by memory, residency, activation. CD127 is known to be enriched on TRM. It is also a marker used in blood to identify regulatory T cells by surface expression. By preventing the enzymatic cleavage of surface markers during tissue digestion, we were able to identify a CD127+FOXP3+ T cell subset. We find its expression is dichotomous with TIGIT expression. The CD127+ Tregs express markers that are associated with TRM. Lastly, we examined the mononuclear phagocyte and CD4+ T cell landscape in human anogenital mucosa by single cell RNA sequencing (sc-RNAseq). We apply scRNA-Seq to conclude that Langerhans cells are not the only cells within the stratified epidermal layer of tissues, as the latter also contain conventional DC2 (cDC2). Further, we examine tissue pDCs and Axl+Siglec6+ DCs (ASDCs) by RNA sequencing and compare these to their counterparts in blood

Examining childhood trauma and comorbid mental disorders among people with opioid use disorder

Opioid use disorder (OUD) is characterised by persistent opioid use despite the experience of harm. Current literature suggests that childhood trauma and mental disorders are risk factors for illicit OUD. However, little is known about the effects of childhood trauma and mental disorders on OUD-related harm. Our understanding of this relationship is also limited by imprecise childhood trauma and mental disorder prevalence data; few studies on people prescribed opioids for chronic non-cancer pain (CNCP); and a lack of research on interventions that reduce harm related to childhood trauma and mental disorders among people with OUD. This thesis uses multiple study methods on data from several different opioid-using populations to broaden our understanding of the relationship between childhood trauma, mental disorders, and OUD. The studies in this thesis aimed to 1) estimate the prevalence of specific childhood trauma types and mental disorders among people with OUD; 2) investigate childhood trauma as an independent risk factor for OUD among a large sample of people prescribed opioids for CNCP; 3) explore the effects of childhood trauma and mental disorders on OAT engagement, crime, and mortality among people with illicit OUD; and 4) examine the associations between OAT and causes of mortality, including suicide, among people with OUD. Multiple key findings emerged from this thesis. The first two studies confirmed that people with OUD experience childhood trauma and mental disorders at far higher rates than the general population. The third study of people prescribed opioids for CNCP found that childhood trauma independently increased the risk of OUD after controlling for mental disorders and other known OUD risk factors. The fourth study found that childhood trauma and most mental disorders were related to poorer OAT engagement and higher criminal offending rates among people with OUD. The fifth study, a systematic review and meta-analysis, found that OAT was associated with reduced risk of multiple causes of mortality, including suicide, among people with OUD. Findings from these studies highlight the importance of addressing childhood trauma and mental disorders in both the treatment of OUD and the management of CNCP with pharmaceutical opioids. The results can inform clinical guidelines, policy, and future research that impact people with OUD and those prescribed opioids for CNCP

Identifying Genes and Novel Variants Involved in Nonsyndromic Hearing Impairment, and Assessment of the Psychosocial Burden of Hearing Impairment in Cameroon

Background Hearing impairment (HI) is the most common sensory disability and occurs in about 1 per 1000 live births in high-income countries, with a much higher incidence of up to 6 per 1000 live births in sub-Saharan Africa (SSA). HI can be due to environmental or genetic causes, and in many cases, it is not possible to establish a definite aetiology. Hereditary HI contributes to 30% to 50% of HI cases in SSA. Hereditary HI can be syndromic or non-syndromic, depending on whether it is associated with additional abnormalities in other organs or not. Non-syndromic HI (NSHI) accounts for 70% of hereditary hearing loss, and is genetically highly heterogeneous, with approximately 170 loci and 121 genes identified to date. Studies in European and Asian populations have identified pathogenic variants in GJB2 (MIM: 121011), and GJB6 (MIM: 604418) genes as the major contributors to autosomal recessive NSHI (ARNSHI). The genetic aetiology of HI in Cameroon is unclear, as previous studies have found no contribution of GJB2 and GJB6 genes to NSHI in Cameroon. However, patients included in those studies consisted of both familial and isolated cases, therefore, underlying environmental/multifactorial causes in some cases cannot be excluded (especially for the isolated cases). Six loci for X-linked HI have been described to date, including DFNX3 (Xp21.2), where DMD is located. Variants in DMD in humans are known to be responsible for Duchenne muscular dystrophy (DMD; MIM: 310200), and Becker muscular dystrophy (BMD; MIM: 300376), an Xlinked recessive disorder. Previous studies have demonstrated that mdx mice, (an animal knockout model for DMD), have an increased threshold for hearing when compared to wildtype mice. However, the contribution of DMD to HI in humans has not been extensively studied. Besides, most of the previous studies on DMD were conducted in Caucasians, Asians, and Arabs; therefore, little is known about the features of this condition in Africans. Parents of children with HI tend to face challenges of parenting especially in terms of communication and social interaction. In Africa, parent's perceived causes of deafness vary from environmental factors to mysterious (“evil forces”) or superstitious beliefs. Also, the attitude of the society towards people with HI does not encourage their participation and involvement in the community, as they face overt discrimination. Aim and methods The aim of this project was to examine the genetic aetiologies of HI in the Cameroonian population, and undercover the challenges faced by persons with HI in Cameroon and their understanding of the causes of HI. This was addressed by 1) Establishing the current status of knowledge on HI in Africa (in terms of prevalence, aetiologies, and genetics aspects) with a particular focus on Cameroon, and assessing the contribution of connexin genes to HI in humans at a global level, through systematic literature reviews; 2) Revisiting the contribution of GJB2 and GJB6 genes to NSHI in 29 multiplex Cameroonian families with NSHI and with strong evidence of non-environmental causes, through targeted gene sequencing and specific multiplex polymerase chain reaction (PCR); 3) Using multiplex ligand-dependent probe amplification (MLPA) technique to investigate the most common variants associated with DMD in Cameroon and assess their possible implication in HI in humans; 4) Performing whole exome sequencing (WES) on 2 Cameroonian multiplex families with NSHI and who tested negative for pathogenic variants in GJB2 and GJB6, to identify the underlying causative genes; 5) Performing in-depth interviews to gain an understanding of the challenges faced by people with HI in Cameroon, their understanding of the causes of hearing impairment (HI), and how challenges could be remedied to improve the quality of life of persons with HI. Results Literature reviews Our first systematic review showed that HI is a public health issue in Cameroon, especially in the elder population where the prevalence of HI is 14.8% in people aged 50 years and more. Environmental factors, including meningitis, impacted wax, and age-related disorders are the leading aetiologies of HI in Cameroon as in many other SSA countries, contributing 52.6% to 62.2% of HI cases. Hereditary HI comprises 0.8% to 14.8% of all cases in Cameroon, and in 32.6% to 37% of HI cases, the origin remains unknown. This contrasts with findings from highincome countries where hereditary HI constitutes the main aetiology of HI, contributing to approximately 50% of cases. NSHI is the most frequent clinical entity and accounts for 86.1% to 92.5% of cases of hereditary HI in the Cameroonian population. No pathogenic variant was described in GJB6 gene, and the prevalence of pathogenic variants in GJB2 ranged from 0% to 0.5%. The prevalence of pathogenic variants in other known NSHI genes was with type 2 Waardenburg syndrome, and three cases of type 2 Usher syndrome were identified in one family. By direct gene sequencing of the coding region of GJB2, no variants were found in any of the 29 families with NSHI. Additionally, through a specific multiplex PCR, the GJB6- D3S1830 deletion which contributes to 9.7% of NSHI cases in Europeans was not identified in any of the patients with HI. Subsequently, a total of 17 males with DMD from 14 families were recruited, aged 14 ± 5.1 (8–23) years. The mean age at onset of symptoms was 4.6 ± 1.5 years, and the mean age at diagnosis was 12.1 ± 5.2 years. Proximal muscle weakness was noted in all patients and calf hypertrophy in the large majority of them (88.2%; 15/17). Flexion contractures were particularly frequent on the ankle (85.7%; 12/14). Wasting of the shoulder girdle and thigh muscles was present in 50% (6/12) and 46.2% (6/13) of patients, respectively. No patient presented with HI. The MLPA found that deletions of at least one exon in DMD occurred in 45.5% of patients (5/11), while duplications were observed in 27.3% (3/11). Both variant types were clustered between exons 45 and 50, and the proportion of de novo variant was estimated at 18.2% (2/11). Whole exome sequencing We submitted DNA samples from five members of a multiplex non-consanguineous Cameroonian family segregating prelingual and progressive ARNSHI for WES. We identified novel bi-allelic compound heterozygous pathogenic variants in CLIC5 (MIM: 607293). The variants identified, i.e. the missense [NM_016929.5:c.224T>C; p.(Leu75Pro)] and the splicing (NM_016929.5:c.63+1G>A), were validated using Sanger sequencing in all seven available family members and co-segregated with HI in the three family members with HI. The three affected individuals were compound heterozygous for both variants, and all unaffected individuals were heterozygous for one of the two variants. Both variants classify as pathogenic by the American College of Medical Genetics (ACMG) guidelines for classification of variants and are absent from the genome aggregation database (gnomAD), UK10K, Greater Middle East (GME) database, and the Single Nucleotide Polymorphism Database (dbSNP), as well in 122 healthy controls from Cameroon. We also did not identify these pathogenic variants in 118 unrelated sporadic cases of NSHI from Cameroon. A second multiplex family was also screened through the use of WES, followed by direct Sanger sequencing in additional patients and control participants. We identified a heterozygous novel missense variant [NM_001174116.2:c.918G>T; p.(Gln306His)] in DMXL2 (MIM:612186) which was transmitted in an autosomal dominant manner, and co-segregates with congenital/prelingual profound to total non-syndromic sensorineural HI in a family from Cameroon. The described family showed a variable expressivity of the HI phenotype. The p.(Gln306His) variant which substitutes a highly conserved glutamine residue is predicted deleterious by various bioinformatics tools and is absent from several genome databases including genome aggregation database (gnomAD), and trans-omics for precision medicine (TOPMed) database. This variant was neither found in 121 healthy controls without personal or family history of HI, nor 112 sporadic cases of NSHI from Cameroon. Our study identified novel variants in CLIC5 and DMXL2 in two Cameroonian families, and provided only the second report of variants in these genes worldwide; thus, strengthening the case for these two genes as candidate genes for NSHI in humans. The psychosocial burden of HI We performed in-depth interviews with 10 HI professionals (healthcare workers, and educationists), and 10 persons affected by HI (persons with HI, and caregivers). The results show that in this study population, the cause of HI is attributed to a variety of causes, including genetics, environmental factors, and a spiritual curse. There were reported cases of stigma and discrimination with persons with HI in the Cameroonian population sometimes seen as having a “mental disorder”. Our participants also highlighted the difficulty that persons with HI have in accessing the necessary education and healthcare services, and suggested the need for policymakers and researchers to develop strategies to improve the social integration of persons with HI and their access to basic social services. This includes 1) Increased awareness amongst the general population, 2) the establishment of more special schools, and 3) building and equipping facilities for proper management of HI. Conclusions Our project confirms that variants in GJB2 and GJB6 genes do not contribute significantly to NSHI in the Cameroonian population. Also, variants in DMD that were shown to be associated with an increased hearing threshold in mice, do not seem to be implicated in HI in Cameroon, neither in previous human studies (although they did not objectively assess hearing using standardized testing methods). Despite the first symptoms of DMD occurring in infancy, the diagnosis is frequently made later in adolescence, indicating an underestimation of the number of cases of DMD in Cameroon. Future screening of deletions and duplications in patients from Cameroon should focus on the distal part of the DMD gene. Subsequently, this study successfully identified the candidate genes in two Cameroonian multiplex families with NSHI through the use of WES, and thus highlights the efficacy of next-generation sequencing techniques in resolving HI cases in Cameroonians and in cases where no pathogenic variants are found in common HI-genes. Additionally, our project which confirms that CLIC5 and DMXL2 genes are associated with HI in humans advocate for the inclusion of these two genes in diagnostic gene panels for NSHI in clinical settings. Last, this study shows the difficult social interaction and access to proper management faced by persons with HI in Cameroon, and highlights the need to educate populations on the causes of HI for a better acceptance of persons with HI in the Cameroonian society