| 1. |
- Ahlberg, Viktor, et al.
(författare)
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Innate immune responses induced by the saponin adjuvant Matrix-M in specific pathogen free pigs
- 2017
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Ingår i: Veterinary Research. - : Springer Science and Business Media LLC. - 0928-4249 .- 1297-9716. ; 48
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Tidskriftsartikel (refereegranskat)abstract
- Saponin-based adjuvants have been widely used to enhance humoral and cellular immune responses in many species, but their mode of action is not fully understood. A characterization of the porcine transcriptional response to Matrix-M was performed in vitro using lymphocytes, monocytes or monocyte-derived dendritic cells (MoDCs) and in vivo. The effect of Matrix-M was also evaluated in specific pathogen free (SPF) pigs exposed to conventionally reared pigs. The pro-inflammatory cytokine genes IL1B and CXCL8 were up-regulated in monocytes and lymphocytes after Matrix-M exposure. Matrix-M also induced IL12B, IL17A and IFNG in lymphocytes and IFN-a gene expression in MoDCs. Several genes were indicated as up-regulated by Matrix-M in blood 18 h after injection, of which the genes for IFN-a and TLR2 could be statistically confirmed. Respiratory disease developed in all SPF pigs mixed with conventional pigs within 1-3 days. Two out of four SPF pigs injected with saline prior to contact exposure displayed systemic symptoms that was not recorded for the four pigs administered Matrix-M. Granulocyte counts, serum amyloid A levels and transcription of IL18 and TLR2 coincided with disease progression in the pigs. These results support further evaluation of Matrix-M as a possible enhancer of innate immune responses during critical moments in pig management.
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| 3. |
- Enqvist, Stina, et al.
(författare)
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Germ Line Origin and Somatic Mutations Determine the Target Tissues in Systemic AL-Amyloidosis
- 2007
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Ingår i: PLoS ONE. - : Public Library of Science (PLoS). - 1932-6203. ; 2:10, s. e981-
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Tidskriftsartikel (refereegranskat)abstract
- BACKGROUND: Amyloid is insoluble aggregated proteins deposited in the extra cellular space. About 25 different proteins are known to form amyloid in vivo and are associated with severe diseases such as Alzheimeŕs disease, prion diseases and type-2 diabetes. Light chain (AL) -amyloidosis is unique among amyloid diseases in that the fibril protein, a monoclonal immunoglobulin light chain, varies between individuals and that no two AL-proteins with identical primary structures have been described to date. The variability in tissue distribution of amyloid deposits is considerably larger in systemic AL-amyloidosis than in any other form of amyloidosis. The reason for this variation is believed to be based on the differences in properties of the amyloidogenic immunoglobulin light chain. However, there is presently no known relationship between the structure of an AL-protein and tissue distribution. METHODOLOGY/PRINCIPAL FINDINGS: We compared the pattern of amyloid deposition in four individuals with amyloid protein derived from variable light chain gene O18-O8, the source of a high proportion of amyloidogenic light chains, and in whom all or most of the fibril protein had been determined by amino acid sequencing. In spite of great similarities between the structures of the proteins, there was a pronounced variability in deposition pattern. We also compared the tissue distribution in these four individuals with that of four other patients with AL-amyloid derived from the L2-L16 gene. Although the interindividual variations were pronounced, liver and kidney involvement was much more evident in the latter four. CONCLUSIONS/SIGNIFICANCE: We conclude that although the use of a specific gene influences the tissue distribution of amyloid, each light chain exhibits one or more determinants of organ-specificity, which originate from somatic mutations and post-translational modifications. Eventual identification of such determinants could lead to improved treatment of patients with AL amyloidosis.
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| 4. |
- Gummesson, Christina, et al.
(författare)
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Entrustable professional activities (EPAs) for undergraduate medical education : development and exploration of social validity
- 2023
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Ingår i: BMC Medical Education. - : BioMed Central (BMC). - 1472-6920. ; 23:1
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Tidskriftsartikel (refereegranskat)abstract
- Background: The development of entrustable professional activities (EPAs) as a framework for work-based training and assessment in undergraduate medical education has become popular. EPAs are defined as units of a professional activity requiring adequate knowledge, skills, and attitudes, with a recognized output of professional labor, independently executable within a time frame, observable and measurable in its process and outcome, and reflecting one or more competencies. Before a new framework is implemented in a specific context, it is valuable to explore social validity, that is, the acceptability by relevant stakeholders.Aim: The aim of our work was to define Core EPAs for undergraduate medical education and further explore the social validity of the constructs.Method and material: In a nationwide collaboration, EPAs were developed using a modified Delphi procedure and validated according to EQual by a group consisting of teachers nominated from each of the seven Swedish medical schools, two student representatives, and an educational developer (n = 16). In the next step, social validity was explored in a nationwide survey. The survey introduced the suggested EPAs. For each EPA, the importance of the EPA was rated, as was the rater’s perception of the present graduates’ required level of supervision when performing the activity. Free-text comments were also included and analyzed.Results: Ten Core EPAs were defined and validated. The validation scores for EQual ranged from 4.1 to 4.9. The nationwide survey had 473 responders. All activities were rated as “important” by most responders, ranging from 54 to 96%. When asked how independent current graduates were in performing the ten activities, 6 to 35% reported “independent”. The three themes of the free text comments were: ‘relevant target areas and content’; ‘definition of the activities’; and ‘clinical practice and learning’.Conclusion: Ten Core EPAs were defined and assessed as relevant for Swedish undergraduate medical education. There was a consistent gap between the perceived importance and the certainty that the students could perform these professional activities independently at the time of graduation. These results indicate that the ten EPAs may have a role in undergraduate education by creating clarity for all stakeholders.
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| 7. |
- Hellman, Stina, et al.
(författare)
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Cytokine responses to various larval stages of equine strongyles and modulatory effects of the adjuvant G3 in vitro
- 2020
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Ingår i: Parasite Immunology. - : Wiley. - 0141-9838 .- 1365-3024. ; 43
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Tidskriftsartikel (refereegranskat)abstract
- Aims To generate different larval stages ofStrongylus vulgarisand to study cytokine responses in cultures of eqPBMC exposed to defined larval stages ofS. vulgarisand cyathostomins with the aim to understand the early immune reaction to these parasites. Methods and results EqPBMC were exposed toS. vulgarislarvae (L3, exsheated L3 and L4) and cyathostomin L3 and analysed for cytokine gene expression. Procedures for decontamination, culturing and attenuation of larvae were established. Transcription of IL-4, IL-5 and IL-13 was induced by bothS. vulgarisand cyathostomin L3. Moulting ofS. vulgarisfrom L3 to L4 stage was accompanied by a shift to high expression of IL-5 and IL-9 (exsheated L3 and L4) and IFN-gamma (L4 only). In parallel, the adjuvant G3 modified the cytokine profile induced by both parasites by reducing the expression of IL-4, IL-5 and IL-10 while concomitantly enhancing the expression of IFN-gamma. Conclusion The L4 stage ofS. vulgarisgenerated a cytokine profile different from that induced by the earlier L3 stage ofS. vulgarisand cyathostomins. This diversity depending on the life cycle stage will have implications for the choice of antigen and adjuvant in future vaccine design.
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| 8. |
- Hellman, Stina, et al.
(författare)
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Equine enteroid-derived monolayers recapitulate key features of parasitic intestinal nematode infection
- 2024
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Ingår i: Veterinary research (Print). - : Springer Nature. - 0928-4249 .- 1297-9716. ; 55:1
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Tidskriftsartikel (refereegranskat)abstract
- Stem cell-derived organoid cultures have emerged as attractive experimental models for infection biology research regarding various types of gastro-intestinal pathogens and host species. However, the large size of infectious nematode larvae and the closed structure of 3-dimensional organoids often hinder studies of the natural route of infection. To enable easy administration to the apical surface of the epithelium, organoids from the equine small intestine, i.e. enteroids, were used in the present study to establish epithelial monolayer cultures. These monolayers were functionally tested by stimulation with IL-4 and IL-13, and/or exposure to infectious stage larvae of the equine nematodes Parascaris univalens, cyathostominae and/or Strongylus vulgaris. Effects were recorded using transcriptional analysis combined with histochemistry, immunofluorescence-, live-cell- and scanning electron microscopy. These analyses revealed heterogeneous monolayers containing both immature and differentiated cells including tuft cells and mucus-producing goblet cells. Stimulation with IL-4/IL-13 increased tuft- and goblet cell differentiation as demonstrated by the expression of DCLK1 and MUC2. In these cytokine-primed monolayers, the expression of MUC2 was further promoted by co-culture with P. univalens. Moreover, live-cell imaging revealed morphological alterations of the epithelial cells following exposure to larvae even in the absence of cytokine stimulation. Thus, the present work describes the design, characterization and usability of an experimental model representing the equine nematode-infected small intestinal epithelium. The presence of tuft cells and goblet cells whose mucus production is affected by Th2 cytokines and/or the presence of larvae opens up for mechanistic studies of the physical interactions between nematodes and the equine intestinal mucosa.
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| 9. |
- Hellman, Stina
(författare)
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Generation of equine enteroids and enteroid-derived 2D monolayers that are responsive to microbial mimics
- 2021
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Ingår i: Veterinary Research. - : Springer Science and Business Media LLC. - 0928-4249 .- 1297-9716. ; 52
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Tidskriftsartikel (refereegranskat)abstract
- Enteroid cultures are three-dimensional in vitro models that reflect the cellular composition and architecture of the small intestine. One limitation with the enteroid conformation is the enclosed lumen, making it difficult to expose the apical surface of the epithelium to experimental treatments. The present study was therefore conducted to generate cultures of equine enteroids and to develop methods for culture of enteroid-derived cells on a two-dimensional plane, enabling easy access to the apical surface of the epithelium. Equine enteroids were established from small intestinal crypts within 7-9 days of culture. Transcriptional analysis of cell type markers confirmed the presence of enterocytes, stem-, Paneth-, proliferative-, enteroendocrine-, goblet- and tuft cells. This cellular composition was maintained over multiple passages, showing that the enteroids can be kept for prolonged periods. The transfer from 3D enteroids to 2D monolayers slightly modified the relative expression levels of the cell type markers, indicating a decrease of goblet- and Paneth cells in the monolayers. Stimulation with the TLR2, 3 and 4 agonists Pam3CSK4, Poly I:C and LPS, respectively, induced the pro-inflammatory cytokines TNF-alpha and IL-8, while the TLR5 agonist FliC only induced TNF-alpha. In addition, an up-regulation of TGF-beta, IL-33 and IFN-beta was recorded after exposure to lipofected Poly I:C that also affected the monolayer integrity. Thus, the equine enteroid-derived 2D monolayers described in the present study show both genetic and functional similarities with the equine intestine making it an interesting in vitro model for studies demanding access to the apical surface, e.g. in studies of host-microbe interactions.
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| 10. |
- Hellman, Stina
(författare)
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Immunological insights into equine responses against Strongylus vulgaris : Ex vivo studies using equine intestinal organoids and blood mononuclear cells
- 2021
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Doktorsavhandling (övrigt vetenskapligt/konstnärligt)abstract
- Anthelminthic drugs have controlled parasitic infections in veterinary species for decades, but today this use is questioned due to concerns about antimicrobial resistance. For most microorganisms, a feasible alternative is vaccination but for parasites, commercial vaccines are scarce. This is partly due to the complex life cycles of most helminths together with their immunomodulatory capacity. In the horse, Strongylus vulgaris has recently gained attention due to its re-emergence in Sweden. The overall aim of the present thesis was therefore to establish ex vivo methods to study responses against S. vulgaris that can guide formulation of a future vaccine. In the search for relevant antigens, protocols for generating defined larval (L) stages were established. These S. vulgaris preparations, alone or in combination with an adjuvant, were evaluated for their cytokine-inducing capacity. All larval preparations induced a Th2 profile in equine PBMC characterised by up-regulation of IL-4, IL-5, IL-9, IL-13 and TSLP. The L4 stage skewed this response by also up-regulating IFN-γ. The presence of a novel saponin adjuvant affected the Th2 profile induced by the L3 stage of S. vulgaris. To study responses to S. vulgaris at the site of infection, equine intestinal stem cells were differentiated into intestinal organoids. From these, monolayer cultures were established that displayed both genetic and functional similarities with the equine intestine, expressing pro-inflammatory cytokine genes at exposure to TLR-agonists. These organoid monolayers were applicable for generation of ex vivo co-cultures with equine PBMC and S. vulgaris larvae. Overall, this thesis provides new insights into the biology of S. vulgaris infection and ex vivo methods that will aid in the development of a future vaccine.
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