Detection of allergen- and mitogen-induced human cytokine transcripts using a competitive polymerase chain reaction

Huang, Shau Ku; Essayan, David M.; Krishnaswamy, Guha; Yi, Ming; Kumai, Megumi; Su, Song Nan; Xiao, Hui; Lawrence, Marcus; Liu, Mark C.

doi:10.1016/0022-1759(94)90052-3

Cited by 39 publications

(10 citation statements)

References 22 publications

Supporting

Mentioning

Contrasting

Order By: Relevance

“…Cytokine gene expression was determined as previously described (20,21). Briefly, either 5 ϫ 10 6 PBMCs/condition or 2 ϫ 10 5 clonal T cells/condition with 3 ϫ 10 6 APCs/ condition were cultured in the absence and presence of antigen in slanted 14-ml polypropylene tubes.…”

Section: Gene Expression Assaysmentioning

confidence: 99%

Differential Regulation of Human, Antigen-specific Th1 and Th2 Responses by the B-7 Homologues, CD80 and CD86

Bashian

Braun

Huang

et al. 1997

Am J Respir Cell Mol Biol

Self Cite

View full text Add to dashboard Cite

A selectivity of B7.1 (CD80) for promoting Th1 responses and B7.2 (CD86) for promoting Th2 responses in the murine system has recently been suggested. The present study explores this hypothesis, using human PBMCs and antigen-specific Th1 and Th2 clones. Proliferative responses of peripheral blood mononuclear cells (PBMCs) from ragweed-allergic, tetanus toxoid-immunized individuals were downregulated by treatment with anti-CD86 in ragweed- and tetanus toxoid-driven cultures (% Inhibition = 55 +/- 4 and 61 +/- 12, respectively; P < 0.03 relative to untreated cultures). Gene expression in PBMCs for interleukin (IL)-4, IL-5, and interferon gamma (IFNgamma), assessed by reverse-transcriptase polymerase chain reaction, was also downregulated by treatment with anti-CD86 in both the ragweed- and tetanus toxoid-driven systems. Neither independent efficacy nor synergy with anti-CD86 was apparent with anti-CD80 treatment; two different anti-CD80 blocking antibodies yielded identical results. Conversely, antigen-specific Th1 and Th2 clones were insensitive to treatment with either anti-CD80, anti-CD86, or a combination of the two. Unaffected parameters included proliferative response (P < 0.14 and 0.33, respectively, for Th1 and Th2), proinflammatory cytokine gene expression, and cytokine protein secretion into culture supernatants (P < 0.44 and 0.16, respectively, for IL-4 and IFNgamma). We conclude that CD86 is the primary B7 signaling homologue in human PBMC responses, and that second signal pathways through the B7 homologues have no effect on phenotypically differentiated T helper cells in humans.

show abstract

Section: Gene Expression Assaysmentioning

confidence: 99%

Differential Regulation of Human, Antigen-specific Th1 and Th2 Responses by the B-7 Homologues, CD80 and CD86

Bashian

Braun

Huang

et al. 1997

Am J Respir Cell Mol Biol

Self Cite

View full text Add to dashboard Cite

show abstract

“…The plasmid vector, pSP64-poly(A), containing this construct was generously provided by Dr S.-K. Huang (Baltimore, MD). To generate an internal standard for IL-13, the plasmid construct was first linearized by digestion with EcoRI, followed by in vitro transcription with SP6 RNA polymerase (Promega Corp., Madison, WI), as described [19]. The IL-13 cRNA standard was purified and, after quantification, reverse transcribed into cDNA as described below.…”

Section: Competitive Rt-pcrmentioning

confidence: 99%

IL-13 over-expression in skin is not confined to IgE-mediated skin inflammation

Ploeg¹,

Tehrani²,

Matuseviciene³

et al. 1997

Clinical and Experimental Immunology

View full text Add to dashboard Cite

SUMMARYIL-13 is produced by T cells and, like IL-4, it can induce the production of IgE and IgG4. In order to investigate if IL-13 is a specific marker for atopic dermatitis (AD), IL-13 gene expression was analysed in chronic lichenified lesions and non-lesional skin of patients with AD, in involved and non-involved skin of patients with psoriasis, in positive tuberculin reactions in non-atopics, and in the skin of healthy control subjects. Patients with AD (n ¼ 9) showed sensitization to common air-borne allergens (positive Phadiatop) and had total serum IgE values in the range from 10 to 4800 kU/l (median 170 kU/l ). Competitive reverse transcription-polymerase chain reaction (RT-PCR) was used to assess IL-13 gene expression in skin biopsy specimens. IL-13 gene expression was markedly higher in chronic lichenified lesions of patients with AD (P < 0 . 01), and in the positive tuberculin reactions (P < 0 . 01; n ¼ 12) than in skin from healthy control subjects (n ¼ 10). However, there was no significant difference in IL-13 gene expression in the skin of patients with psoriasis (n ¼ 10) and that of healthy control subjects. The dermal cell infiltrates were larger and the relative amount of CD3 þ and CD4 þ cells in these infiltrates was higher in the skin of subjects with a positive tuberculin reaction than in lichenified AD skin. However, these differences were not reflected in differences in IL-13 gene expression. Different triggers of IL-13 gene expression may influence the diverse patterns of inflammation seen in different inflammatory skin disorders.

show abstract

“…1.8. (9) RNA was quantitated by optical density readings at 260 nm, and with DipStick (Invitrogen, SanDiego, CA). The integrity of the 28S and 18S bands was determined by electrophoresis in 2% ethidium bromide-stained agarose gel.…”

Section: Characterization Of Parotid Cell Culturesmentioning

confidence: 99%

Interferon-alpha Upregulates Gene Expression of Aquaporin-5 in Human Parotid Glands

Smith¹,

Siddiqui²,

Modica³

et al. 1999

Journal of Interferon & Cytokine Research

View full text Add to dashboard Cite

Aquaporins are a family of homologous membrane proteins that function as highly selective water channels. Aquaporin-5 (AQP5) is uniquely present in lacrimal and salivary glands, where it accounts for normal tear and saliva production. We tested the hypothesis that orally administered human interferon-alpha (HuIFN-alpha) benefits persons with xerostomia by augmenting the production of AQP5 protein by parotid gland epithelium. Cells from three human parotid glands were cultured with and without human lymphoblastoid IFN-alpha, and assayed for AQP5 mRNA levels by reverse transcriptase polymerase chain reaction (RT-PCR), and AQP5 protein levels by Western blot. Intracellular localization of AQP5 protein was done using confocal microscopy. The functional integrity of the glandular tissue was confirmed by RT-PCR analysis of alpha-amylase 1 and basic proline-rich protein transcripts. AQP5 was constitutively expressed in human parotid gland tissue, with AQP5 protein restricted to the plasma membranes and cytoplasmic vesicles of acinar cells. IFN-alpha augmented AQP5 transcription and protein production in a concentration-dependent manner, and increased the size of intensity of staining of AQP5-containing cytoplasmic vesicles in acinar cells. We conclude that IFN-alpha upregulates AQP5 gene expression in human parotid acinar cells in vitro. To our knowledge, this is the first demonstration that IFN-alpha regulates the gene expression of an aquaporin.

show abstract

Detection of allergen- and mitogen-induced human cytokine transcripts using a competitive polymerase chain reaction

Cited by 39 publications

References 22 publications

Differential Regulation of Human, Antigen-specific Th1 and Th2 Responses by the B-7 Homologues, CD80 and CD86

Differential Regulation of Human, Antigen-specific Th1 and Th2 Responses by the B-7 Homologues, CD80 and CD86

IL-13 over-expression in skin is not confined to IgE-mediated skin inflammation

Interferon-alpha Upregulates Gene Expression of Aquaporin-5 in Human Parotid Glands

Contact Info

Product

Resources

About