| 1. |
- Axelsson, Stina, et al.
(författare)
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Cryopreserved peripheral blood mononuclear cells are suitable for the assessment of immunological markers in type 1 diabetic children
- 2008
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Ingår i: Cryobiology. - : Elsevier BV. - 0011-2240 .- 1090-2392. ; 57:3, s. 201-208
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Tidskriftsartikel (refereegranskat)abstract
- Cryopreserved peripheral blood mononuclear cells (PBMC) are commonly used when assessing immune responses in clinical trials, both for practical reasons and to minimize interassay variation, as samples are often collected and studied over time. This study investigated the effect of cryopreservation on cytokine and chemokine secretion, and on expression of regulatory T-cell associated markers, in samples from children with type 1 diabetes. PBMC were cultured before and after cryopreservation either with GAD(65) or PHA. Secretion of cytokines (IL-5, -6, -10, -12, -13 -17, IFN-gamma and TNF-alpha) and chemokines (IP-10, MCP-1, MIP-1 alpha, MIP-1 beta and RANTES) was analysed in cell supernatants using multiplex fluorochrome technique (Luminex). Expression of FOXP3 and TGF-beta mRNA was detected by multiplex real-time RT-PCR. Increased spontaneous secretion of IL-6, -10, -12, -13, IFN-gamma and MCP-1, and mRNA expression of FOXP3 and TGF-beta, was detected after cryopreservation. Stimulation with GAD65 induced higher levels of IL-6, IFN-gamma, TNF-alpha and MIP-1 alpha, whereas lower secretion was found for IL-10 and IL-13 in cryopreserved PBMC. Stimulation with PHA induced lower secretion of IP-10, MCPA and RANTES and FOXP3 mRNA expression after cryopreservation. Thus, cryopreserved PBMC were suitable to assess the immunological markers included in this study, even though their expression could differ from freshly handled cells.
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| 2. |
- Diaz Cruz, Maria Araceli, et al.
(författare)
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Characterization of methylation patterns associated with lifestyle factors and vitamin D supplementation in a healthy elderly cohort from Southwest Sweden
- 2022
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Ingår i: Scientific Reports. - : Nature Portfolio. - 2045-2322. ; 12:1
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Tidskriftsartikel (refereegranskat)abstract
- Numerous studies have shown that lifestyle factors, such as regular physical activity and vitamin D intake, may remarkably improve overall health and mental wellbeing. This is especially important in older adults whose vitamin D deficiency occurs with a high prevalence. This study aimed to examine the influence of lifestyle and vitamin D on global DNA methylation patterns in an elderly cohort in Southwest of Sweden. We also sought to examine the methylation levels of specific genes involved in vitamin D's molecular and metabolic activated pathways. We performed a genome wide methylation analysis, using Illumina Infinium DNA Methylation EPIC 850kBeadChip array, on 277 healthy individuals from Southwest Sweden at the age of 70-95. The study participants also answered queries on lifestyle, vitamin intake, heart medication, and estimated health. Vitamin D intake did not in general affect methylation patterns, which is in concert with other studies. However, when comparing the group of individuals taking vitamin supplements, including vitamin D, with those not taking supplements, a difference in methylation in the solute carrier family 25 (SCL25A24) gene was found. This confirms a previous finding, where changes in expression of SLC25A24 were associated with vitamin D treatment in human monocytes. The combination of vitamin D intake and high physical activity increased methylation of genes linked to regulation of vitamin D receptor pathway, the Wnt pathway and general cancer processes. To our knowledge, this is the first study detecting epigenetic markers associated with the combined effects of vitamin D supplementation and high physical activity. These results deserve to be further investigated in an extended, interventional study cohort, where also the levels of 25(OH)D3 can be monitored.
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| 3. |
- Diaz Cruz, Maria Araceli, et al.
(författare)
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Cis-regulatory elements in conserved non-coding sequences of nuclear receptor genes indicate for crosstalk between endocrine systems
- 2021
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Ingår i: Open Medicine (Poland). - : De Gruyter Open. - 2391-5463. ; 16:1, s. 640-650
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Tidskriftsartikel (refereegranskat)abstract
- Nuclear receptors (NRs) are ligand-activated transcription factors that regulate gene expression when bound to specific DNA sequences. Crosstalk between steroid NR systems has been studied for understanding the development of hormone-driven cancers but not to an extent at a genetic level. This study aimed to investigate crosstalk between steroid NRs in conserved intron and exon sequences, with a focus on steroid NRs involved in prostate cancer etiology. For this purpose, we evaluated conserved intron and exon sequences among all 49 members of the NR Superfamily (NRS) and their relevance as regulatory sequences and NR-binding sequences. Sequence conservation was found to be higher in the first intron (35%), when compared with downstream introns. Seventy-nine percent of the conserved regions in the NRS contained putative transcription factor binding sites (TFBS) and a large fraction of these sequences contained splicing sites (SS). Analysis of transcription factors binding to putative intronic and exonic TFBS revealed that 5 and 16%, respectively, were NRs. The present study suggests crosstalk between steroid NRs, e.g., vitamin D, estrogen, progesterone, and retinoic acid endocrine systems, through cis-regulatory elements in conserved sequences of introns and exons. This investigation gives evidence for crosstalk between steroid hormones and contributes to novel targets for steroid NR regulation.
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| 4. |
- Diaz Cruz, Maria Araceli, et al.
(författare)
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Differential expression of protein disulfide-isomerase A3 isoforms, PDIA3 and PDIA3N, in human prostate cancer cell lines representing different stages of prostate cancer
- 2021
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Ingår i: Molecular Biology Reports. - : Springer. - 0301-4851 .- 1573-4978. ; 48:3, s. 2429-2436
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Tidskriftsartikel (refereegranskat)abstract
- Prostate cancer (PCa) is a highly heterogeneous and unpredictable progressive disease. Sensitivity of PCa cells to androgens play a central role in tumor aggressiveness but biomarkers with high sensitivity and specificity that follow the progression of the disease has not yet been verified. The vitamin D endocrine system and its receptors, the Vitamin D Receptor (VDR) and the Protein Disulfide-Isomerase A3 (PDIA3), are related to anti-tumoral effects as well as carcinogenesis and have therefore been suggested as potential candidates for the prevention and therapy of several cancer forms, including PCa. In this study, we evaluated the mRNA expression of VDR and PDIA3 involved in vitamin D signaling in cell lines representing different stages of PCa (PNT2, P4E6, LNCaP, DU145 and PC3). This study further aimed to evaluate vitamin D receptors and their isoforms as potential markers for clinical diagnosis of PCa. A novel transcript isoform of PDIA3 (PDIA3N) was identified and found to be expressed in all PCa cell lines analyzed. Androgen-independent cell lines showed a higher mRNA expression ratio between PDIA3N/PDIA3 contrary to androgen-dependent cell lines that showed a lower mRNA expression ratio between PDIA3N/PDIA3. The structure of PDIA3N differed from PDIA3. PDIA3N was found to be a N-truncated isoform of PDIA3 and differences in protein structure suggests an altered protein function i.e. cell location, thioredoxin activity and affinity for 1,25(OH)2D3. Collectively, PDIA3 transcript isoforms, the ratio between PDIA3N/PDIA3 and especially PDIA3N, are proposed as candidate markers for future studies with different stages of PCa progression.
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| 5. |
- Diaz Cruz, Maria Araceli
(författare)
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Exploring vitamin D and steroid hormone receptors – from healthy elderly to prostate cancer cells
- 2022
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Doktorsavhandling (övrigt vetenskapligt/konstnärligt)abstract
- The genetic background together with environmental factors and lifestyle are key contributors to the health of an individual. Genetic background is inherited and irreversible unless mutations occur. However, lifestyle habits (i.e., diet, stress, physical activity, smoking, and alcohol consumption) are modifiable factors that contribute to health or disease by affecting methylation of DNA, which regulates transcription of genes.One of the most relevant lifestyle habits for health is maintaining adequate vitamin D levels in the body as vitamin D promotes calcium and phosphate absorption, supports the nervous and immune system function, and protects bone and muscle structure. Extreme low levels of vitamin D, vitamin D deficiency, has become a global public health concern, especially in the elderly population as vitamin D deficiency can lead to several health problems such as bone fracture, decreased muscle strength, cardiovascular and autoimmune diseases, depression, and breast, pancreatic, and prostate cancer.Prostate cancer is an uncontrolled growth of cells within the prostate gland in the male reproductive system. Human prostate carcinomas are sensitive to androgens, and hormonal ablation therapy gives a temporary remission, followed by a relapse to an androgen-insensitive state. This indicates that steroid hormones, especially androgens, play a significant role in human prostatic carcinogenesis. The molecular effect of vitamin D as a steroid hormone and which steroid hormone receptor (SHR) mediates this effect are not fully understood.This research project aims to increase our knowledge about SHRs, primarily the vitamin D receptors, in both health and disease, focusing on genomic, epigenomic, and transcriptomic perspectives in healthy elderly individuals and prostate cancer cells.The results from the studies in this thesis could help us understand the importance of a healthy lifestyle, which includes vitamin D for health, where we found specific methylation markers involved in the down-regulation of cancer pathways that are associated with high physical activity and vitamin D supplementation. We have further confirmed that SHRs rarely work in isolation but rather as a crosstalk at the genomic level to regulate their transcription. Hopefully, this will help clarify the modulation of transcriptional responses in SHRs and explain the development of steroid hormone-dependent cancers such as prostate cancer. Last, but not least, we revealed that genetic and transcriptional markers are associated with the putative vitamin D receptor the protein disulfide isomerase family A member 3 (PDIA3). The genetic markers were detected in a healthy elderly population under vitamin D supplementation. The transcriptional markers, PDIA3, and a novel discovered isoform of PDIA3 (PDIA3N) were related to the androgen and cancer stage of prostate cancer cells and therefore are proposed as candidate markers for clinical diagnosis of prostate cancer.Altogether, these findings support the relevance of studying vitamin D and steroid hormone receptors, especially the PDIA3 receptor, to understand some of the factors related to healthy aging and the etiology and progression of prostate cancer.
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| 6. |
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| 7. |
- Hedman Hjorth, Maria, et al.
(författare)
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Impaired CD4+ and CD8+ T cell phenotype and reduced chemokine secretion in recent-onset type 1 diabetic children
- 2008
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Ingår i: Clinical and Experimental Immunology. - : Oxford University Press (OUP). - 0009-9104 .- 1365-2249. ; 153:3, s. 360-368
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Tidskriftsartikel (refereegranskat)abstract
- Although the role of the T cell-mediated autoimmune reaction in type 1 diabetes (T1D) is conclusive, studies including data from human circulating CD4+ and CD8+ lymphocytes subsets during the disease onset and posterior development are scarce. Further, chemokines and chemokine receptors are key players in the migration of pathogenic T cells into the islets of NOD mice developing T1D, but few studies have investigated these markers in human T1D patients. We studied the expression of T helper 1 (Th1) and Th2 associated chemokine receptors, and the two isoforms of CD45 leukocyte antigen on CD4+ and CD8+ lymphocytes from T1D and healthy children, as well as the secretion of chemokines in cell supernatants in peripheral blood mononuclear cells. Our results showed increased expression of CCR7 and CD45RA, and reduced CD45RO on CD8+ cells among recent-onset T1D patients. The percentages of CD4+ cells expressing CXC chemokine receptor 3 (CXCR3), CXCR6 and CCR5, and the secretion of interferon-γ-induced protein-10, monocyte chemoattractant protein-1, macrophage inflammatory protein (MIP)-1a and MIP-1β was lower among diabetics. Low expression of Th1-associated receptors and secretion of chemokines, together with an increased amount of CD8+ cells expressing CD45RA and CCR7 in T1D patients therefore might represent suboptimal Th function in T1D, leading to impaired T cytotoxic (Tc) responses or alternatively reflect a selective recruitment of Th1 cells into the pancreas.
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| 8. |
- Hedman Hjorth, Maria, 1978-, et al.
(författare)
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Nicotinamide reduces high secretion of IFN-γ in high-risk relatives even though it does not prevent type 1 diabetes
- 2006
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Ingår i: Journal of Interferon and Cytokine Research. - : Mary Ann Liebert Inc. - 1079-9907 .- 1557-7465. ; 26:4, s. 207-213
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Tidskriftsartikel (refereegranskat)abstract
- Type 1 diabetes (T1D) is an autoimmune disease suggested to be of a T helper (Th)1-like origin. This study aimed to investigate the Th1-like and Th2-like profile in high-risk individuals during the prediabetic phase and the immunologic effect of treatment with nicotinamide. High-risk first-degree relatives of T1D patients participating in the European Nicotinamide Diabetes Intervention Trial (ENDIT) were treated with either nicotinamide or placebo. Peripheral blood mononuclear cells (PBMC) were obtained during the prediabetic phase and close to the onset of manifest T1D and from nondiabetic high-risk individuals. Using the sensitive enzyme-linked immunospot (ELISPOT) technique to distinguish Th1-like from Th2-like lymphocytes, secretion of interferon-γ (IFN-γ) and interleukin-4 (IL-4) was analyzed from PBMCs spontaneously and after in vitro stimulation with the diabetes-associated autoantigens, glutamic acid decarboxylase 65 (GAD65, protein and peptide, aa 247-279), recombinant tyrosine phosphatase (IA-2), and heat shock protein (HSP, aa 437-460). High-risk individuals showed high spontaneous as well as autoantigen-induced IFN-γ secretion. Secretion of IFN-γ and the IFN-γ/IL-4 ratio, induced by autoantigens, decreased in individuals developing T1D (p < 0.05), whereas nondiabetic individuals showed an increased IL-4 response (p < 0.05). Thus, a Th1-dominated cytokine profile observed in high-risk individuals inclined toward a diagnosis of diabetes. Nicotinamide caused decreased spontaneous (p = 0.05) and in vitro autoantigen-induced IFN-γ secretion (p < 0.05) and may play a role in immune regulation, even though it has not been shown to prevent T1D. © Mary Ann Liebert, Inc.
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| 9. |
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| 10. |
- Hjorth, Maria, et al.
(författare)
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GAD-alum treatment induces GAD65-specific CD4+CD25highFOXP3+ cells in type 1 diabetic patients.
- 2011
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Ingår i: Clinical Immunology. - : Elsevier. - 1521-6616 .- 1521-7035. ; 138:1, s. 117-126
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Tidskriftsartikel (refereegranskat)abstract
- Type 1 diabetes results from autoimmune destruction of insulin producing pancreatic β-cells. We have shown that treatment with alum-formulated glutamic acid decarboxylase 65 (GAD-alum) preserved residual insulin secretion and induced antigen-specific responses in children with recent onset type 1 diabetes. The aim of this study was to further investigate the immunomodulatory effect of GAD-alum, focusing on CD4+CD25high cells and their association to cytokine secretion. Samples obtained 21 and 30 months after the initial injection of GAD-alum or placebo were included in the present study. GAD65-stimulation enhanced the percentage of CD4+CD25highFOXP3+ cells, but reduced the percentage of CD4+CD25+ cells, in samples from the GAD-alum treated group. Further, the GAD65-induced secretion of IL-5, -10, and -13 correlated with the expression of CD4+CD25highFOXP3+ cells, but inversely with CD4+CD25+ cells. These new data suggest that GAD-alum treatment induced GAD65-specific T cells with regulatory features.
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