| 1. |
- Carlsson, Björn, et al.
(författare)
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Simultaneous generation of cytomegalovirus-specific CD8+ and CD4+ T lymphocytes by use of dendritic cells comodified with pp65 mRNA and pp65 protein
- 2005
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Ingår i: Journal of Infectious Diseases. - : Oxford University Press (OUP). - 0022-1899 .- 1537-6613. ; 192:11, s. 1912-20
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Tidskriftsartikel (refereegranskat)abstract
- Cytomegalovirus (CMV) disease remains a severe complication in patients who have undergone transplantation. Viremia can be prevented and treated by the adoptive transfer of donor-derived CMV-directed T cells. To ensure long-term protection against CMV disease, it is important to transfer CMV antigen-specific T cells that represent both the CD8+ and the CD4+ subsets. In the present study, we used as stimulators dendritic cells (DCs) that were electroporated with in vitro-transcribed 5'-capped polyadenylated messenger RNA (mRNA) that encoded the CMV pp65 protein (i.e., pp65 mRNA). These DCs could efficiently activate CMV-directed CD8+ T cells, as assayed by tetramer staining, interferon- gamma production, and cytolytic activity. We also used DCs that were pulsed with a recombinant pp65 protein to activate CMV-directed CD4+ T cells. When DCs were comodified with pp65 mRNA and pp65 protein, large numbers of CMV-directed CD8+ and CD4+ T cells were generated simultaneously. The approach outlined in the present study can be adapted for a clinical protocol that circumvents potential virus-related biohazards and is available to all patients independently of their human leukocyte antigen haplotype.
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| 2. |
- Hou, Mingyan, et al.
(författare)
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Capsaicin receptor immunoreactivity in the human trigeminal ganglion.
- 2002
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Ingår i: Neuroscience Letters. - 0304-3940. ; 330:3, s. 223-226
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Tidskriftsartikel (refereegranskat)abstract
- The cloned capsaicin receptor, also known as vanilloid receptor subtype 1 (VR1) receptor, has been demonstrated to be an integral membrane protein with homology to a family of putative store-operated calcium channels. The VR1 receptor is activated not only by capsaicin but also by noxious heat and protons, and therefore it is suggested as a molecular integrator of chemical and physical stimuli that elicit pain. In the present study, indirect immunofluorescence detected a small number of neurons that are VR1 receptor immunoreactive (ir) (171 versus 1038 or 16% of all neuronal cell bodies) in the human trigeminal ganglion (TG). In addition, RT-PCR confirmed the presence of VR1 mRNA in the human TG. It has been hypothesized that TG neuronal cell bodies are the source of capsaicin-stimulated release of calcitonin gene-related peptide (CGRP), and hence co-localization experiments were performed. Around 10% of the VR1 receptor-ir is expressed on neurons that contain CGRP-ir (ten among 74) in the human TG, indicating that capsaicin may act through the VR1 receptor to modulate the release of CGRP and in turn to modulate pain. We observed that 8% of the VR1 receptor-ir neuronal cell bodies contain substance P-ir and 5% nitric oxide synthase. Capsaicin can release nitric oxide, CGRP and substance P from sensory nerves and contribute to central sensitization.
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| 3. |
- Hou, Mingyan, et al.
(författare)
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Increase in cardiac P2X1-and P2Y2-receptor mRNA levels in congestive heart failure
- 1999
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Ingår i: Life Sciences. - 1879-0631. ; 65:11, s. 1195-1206
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Tidskriftsartikel (refereegranskat)abstract
- We wanted to study the expression of P2-receptors at the mRNA-level in the heart and if it is affected by congestive heart failure (CHF). To quantify the P2 receptor mRNA-expression we used a competitive RT-PCR protocol which is based on an internal RNA standard. The P2 receptor mRNA-expression was quantified in hearts from CHF rats and compared to sham-operated rats. Furthermore, the presence of receptor mRNA was studied in the myocardium from patients with heart failure. In the sham operated rats the G-protein coupled P2Y-receptors were expressed at a higher level than the ligand gated ion-channel receptor (P2X1). Among the P2Y-receptors the P2Y6-receptor was most abundantly expressed (P2Y6 > P2Y1 > P2Y2 = P2Y4 > P2X1). A prominent change was seen for the P2X1- and P2Y2-receptor mRNA levels which were increased 2.7-fold and 4.7-fold respectively in the myocardium from the left ventricle of CHF-rats. In contrast, the P2Y1-, P2Y4- and P2Y6-receptor mRNA levels were not significantly altered in CHF rats. In human myocard the P2X1-, P2Y1-, P2Y2-, P2Y6- and P2Y11-receptors were detected by RT-PCR in both right and left atria and ventricles, while the P2Y4-receptor band was weak or absent. In conclusion, most of the studied P2-receptors were expressed in both rat and human hearts. Furthermore, the P2X1- and P2Y2-receptor mRNA were upregulated in CHF, suggesting a pathophysiological role for these receptors in the development of heart failure.
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| 4. |
- Hou, Mingyan, et al.
(författare)
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MAPKK-dependent growth factor-induced upregulation of P2Y2 receptors in vascular smooth muscle cells
- 1999
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Ingår i: Biochemical and Biophysical Research Communications. - : Elsevier BV. - 1090-2104 .- 0006-291X. ; 258:3, s. 648-652
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Tidskriftsartikel (refereegranskat)abstract
- The ATP- and UTP-sensitive P2Y2 receptor which mediates both contractile and mitogenic effects has recently been shown to be upregulated in the synthetic phenotype of the vascular smooth muscle cell (VSMC). Using a competitive RT-PCR we demonstrate that the P2Y2 receptor mRNA is increased by fetal calf serum and other growth factors in a MAPKK-dependent way. This was confirmed at the functional level by examining UTP-stimulated release of intracellular Ca2+. Furthermore, the P2Y2 receptor mRNA is positively autoregulated by ATP and the mRNA is rapidly degraded with only 26% remaining after 1 h in the presence of actinomycin D. Our results indicate growth factor regulation and rapid turnover of the P2Y2 receptor mRNA, which may be of importance in atherosclerosis and neointima formation after balloon angioplasty.
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| 5. |
- Hou, Mingyan
(författare)
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Messenger molecules and receptors in the human trigeminovascular system
- 2001
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Doktorsavhandling (övrigt vetenskapligt/konstnärligt)abstract
- The initiation of a migraine attack involves a primary central nervous system dysfunction with subsequent activation of the trigeminovascular system. In trigeminal ganglia, the expression of calcitonin gene-related peptide (CGRP), substance P (SP) and nitric oxide synthase (NOS) have been demonstrated by immunocytochemistry. During migraine attacks, the level of released CGRP is increased. Meanwhile, clinical trials have shown that 5-HT receptor agonists can alleviate headache by the inhibition of the release of CGRP from the sensory nervous system. Although there is no evidence of increased levels of SP or NO during migraine, the involvement of SP and NO in migraine has been suggested by different ways. A NOS inhibitor inhibits migraine while a NO donor causes migraine. While SP is not involved in early attacks, it may come later and be responsible for neurogenic inflammation. In this thesis, the aims were to study the distribution of 5-HT1B and 5-HT1D receptors, the capsaicin receptor and nociceptin as well as their co-localization with CGRP, SP or NOS. All the studied receptors and nociceptin were localized in the human trigeminal ganglion and they were mainly present in medium-sized neuronal cell bodies. The existence of the capsaicin receptor and the nociceptin receptor in addition has also been studied at the mRNA level in human trigeminal ganglia and in cerebral arteries by RT-PCR. They existed in human trigeminal ganglia but not in human cerebral arteries. This was further confirmed by in vitro pharmacology, which showed no vasomotor effect in human cerebral arteries by nociceptin. All the studied receptors and nociceptin were co-localized with CGRP, SP and NOS. In this thesis, we also characterized the vasoconstrictory effect of the P2Y6 receptor in human cerebral arteries by in vitro pharmacology. The RT-PCR was used to confirm the existence of P2Y6 receptors in the human cerebral artery, putatively this may indicate a new anti-migraine mechanism. Further study detected that P2Y6 receptors can induce proliferation of vascular smooth muscle cells, via phospholipase C, protein kinase C delta and a tyrosine kinase pathway. Quantitative competitive RT-PCR revealed that P2Y6 receptor mRNA is rapid turnover and is regulated by growth factors that can be released under physiological and pathophysiological conditions.
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| 6. |
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| 7. |
- Hou, Mingyan, et al.
(författare)
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UDP acts as a growth factor for vascular smooth muscle cells by activation of P2Y(6) receptors.
- 2002
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Ingår i: American Journal of Physiology: Heart and Circulatory Physiology. - : American Physiological Society. - 1522-1539 .- 0363-6135. ; 282:2, s. 784-792
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Tidskriftsartikel (refereegranskat)abstract
- Mitogenic effects of the extracellular nucleotides ATP and UTP are mediated by P2Y(1), P2Y(2), and P2Y(4) receptors. However, it has not been possible to examine the highly expressed UDP-sensitive P2Y(6) receptor because of the lack of stable, selective agonists. In rat aorta smooth muscle cells (vascular smooth muscle cells; VSMC), UDP and UTP stimulated (3)H-labeled thymidine incorporation with similar pEC(50) values (5.96 and 5.69). Addition of hexokinase did not reduce the mitogenic effect of UDP. In cells transfected with P2Y receptors the stable pyrimidine agonist uridine 5'-O-(2-thiodiphosphate) (UDPbetaS) was specific for P2Y(6) with no effect on P2Y(1), P2Y(2), or P2Y(4) receptors. UDPbetaS stimulated [(3)H]thymidine and [(3)H]leucine incorporation and increased cell number in VSMC. Flow cytometry demonstrated that UDP stimulated cell cycle progression to both the S and G(2) phases. The intracellular signal pathways were dependent on phospholipase C, possibly protein kinase C-delta, and a tyrosine kinase pathway but independent of G(i) proteins, eicosanoids, and protein kinase A. The half-life of P2Y(6) receptor mRNA was <1 h by competitive RT-PCR. The mitogen-activated protein kinase kinase inhibitor PD-098059 significantly suppressed, whereas ATP and interleukin-1beta upregulated, expression of P2Y(6) receptor mRNA. The results demonstrate that UDP stimulates mitogenesis through activation of P2Y(6) receptors and that the receptor is regulated by factors important in the development of vascular disease.
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| 8. |
- Malmsjö, Malin, et al.
(författare)
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Potent P2Y6 receptor mediated contractions in human cerebral arteries
- 2003
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Ingår i: BMC Pharmacology. - : Springer Science and Business Media LLC. - 1471-2210. ; 3
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Tidskriftsartikel (refereegranskat)abstract
- BACKGROUND: Extracellular nucleotides play an important role in the regulation of vascular tone and may be involved in cerebral vasospasm after subarachnoidal haemorrhage. This study was designed to characterise the contractile P2 receptors in endothelium-denuded human cerebral and omental arteries. The isometric tension of isolated vessel segments was recorded in vitro. P2 receptor mRNA expression was examined by RT-PCR. RESULTS: In human cerebral arteries, the selective P2Y6 receptor agonist, UDPbetaS was the most potent of all the agonists tested (pEC50 = 6.8 PlusMinus; 0.7). The agonist potency; UDPbetaS > alphabeta-MeATP > UTPgammaS > ATPgammaS > ADPbetaS = 0, indicated the presence of contractile P2X1 P2Y2, P2Y4 and P2Y6, but not P2Y1 receptors, in human cerebral arteries. In human omental arteries, UDPbetaS was inactive. The agonist potency; alphabeta-MeATP > ATPgammaS = UTPgammaS > ADPbetaS = UDPbetaS = 0, indicated the presence of contractile P2X1, and P2Y2 receptors, but not P2Y1 or P2Y6 receptors, in human omental arteries. RT-PCR analysis of endothelium-denuded human cerebral and omental arteries demonstrated P2X1, P2Y1, P2Y2 and P2Y6 receptor mRNA expression. There were no bands for the P2Y4 receptor mRNA in the omental arteries, while barely detectable in the cerebral arteries. CONCLUSIONS: P2Y6 receptors play a prominent role in mediating contraction of human cerebral arteries. Conversely, no such effect can be observed in human omental arteries and previous results confirm the absence of P2Y6 receptors in human coronary arteries. The P2Y6 receptor might be a suitable target for the treatment of cerebral vasospasm.
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| 9. |
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