RESUMO
BACKGROUND: Chronic primary pain (CPP) as a diagnosis has been introduced in the recent International Classification of Diseases, 11th Revision (ICD-11). CPP captures the experience of pain as the primary problem, without an underlying attributable cause. Dissemination of UK guidance regarding CPP represents the first time it has been recognised as a condition in its own right. Little is known regarding General Practitioner (GP) views concerning caring for patients with CPP and how related guidance is viewed and applied in practice. AIM: To explore GP perspectives in relation to caring for people with CPP, including challenges encountered and use of related guidelines in practice. DESIGN & SETTING: A UK-wide qualitative interview study in primary care. METHOD: Purposive and snowball sampling were used to recruit 15 GP participants from England, Northern Ireland, Wales and Scotland. Semi-structured interviews were undertaken and analysed using reflexive thematic analysis. RESULTS: Three main themes were generated: (1) "How to start? Problematic beginnings" referred to difficulties regarding diagnosis; (2) "Where to go? Mapping the management challenge" and (3) "How to get there? Navigating strategies and response", explored GP awareness and acceptability of UK guidelines for chronic pain. Areas identified for potential improvement included increased access to NPM and secondary care services, support with de-prescribing and an expanded multidisciplinary team input. CONCLUSION: CPP is complex to both diagnose and manage. Although guidelines provide a useful framework, they pose challenges when translating into day-to-day practice.
RESUMO
Molluscum contagiosum virus (MCV), a poxvirus pathogenic for humans, replicates well in human skin in vivo, but not in vitro in standard monolayer cell cultures. In order to determine the nature of the replication deficiency in vitro, the MCV infection process in standard culture has to be studied step by step. The method described in this chapter uses luciferase and GFP reporter constructs to measure poxviral mRNA transcription activity in cells in standard culture infected with known quantities of MCV or vaccinia virus. Briefly, MCV isolated from human tissue specimen is quantitated by PCR and used to infect human HEK293 cells, selected for ease of transfection. The cells are subsequently transfected with a reporter plasmid encoding firefly luciferase gene under the control of a synthetic early/late poxviral promoter and a control plasmid encoding a renilla luciferase reporter under the control of a eukaryotic promoter. After 16 h, cells are harvested and tested for expression of luciferase. MCV genome units are quantitated by PCR targeting a genome area conserved between MCV and vaccinia virus. Using a GFP reporter plasmid, this method can be further used to infect a series of epithelial and fibroblast-type cell lines of human and animal origin to microscopically visualize MCV-infected cells, to assess late promoter activation, and, using these parameters, to optimize MCV infectivity and gene expression in more complex eukaryotic cell culture models.
